-^ ;UMUlttUlUHiWitUii,i CALIFCRNIA COLLEGE ^J- •• APR fj nn IBVIWI, CWJfORNlA 92GG4 HEMATOLOGICAL ATLAS ■ WITH A DESCRIPTION OF THE TECHNIC OF BLOOD EXAMINATION BY ' / V Priv. Doz. Dr. Karl Schleip Scientific Assistant in the Medical Clinic University of Freiburg i/ B. ENGLISH ADAPTATION OF TEXT BY Frederic E. Sondern, M.D. Professor of Clinical Pathology New York Post-Graduate Medical School and Hospital. Director of Clinical Laboratory New York Lying-in Hospital Witt) 71 Colorcb aUuStrationsf REVISED AND IMPROVED EDITION NEW YORK. REBMAN COMPANY mo Copyright, 1920, nv BEBMAN COMPANY New York Printed in America A PREFACE TO THE ENGLISH EDITION The paucity of good pictures illustrating the changes in the microscopic appearance of the blood which have become apparent by the use of panoptic staining methods is e\'ident to the teacher of Clinical Hematology. To this fact noted by the author, and to the opportunities offered him by the hematological ^ material available at the Medical CUnic of the Uni- versify of Freiburg, this Atlas owes its origin. An accurate picture of characteristic changes is often better than a detailed description or an imperfect specimen. The object of the Atlas is not only to assist in teaching the subject but also to serve as a guide for the private study of the clinician; and the brief diag- j' nostic points and explanatory comments may be of J\ value in this connection. In the nomenclature of the blood cells the universally used terms have been se- n lected, and those based on theories not yet firmly '* ■ established have been avoided. A sufficiently detailed technic of the cHnical methods used in blood examination has been in- ^ eluded, with the hope of enhancing the usefulness of the book, and the compMcated procedures demanding extensive apparatus and not suited to the use of the chnical worker have been omitted. The original illustrations represent specimens stained with the Leish7nan modification of the Romanowsky method, +- and uniform magnifications of 330 and 750 only have been used. S" Few slight changes have been made in the Enghsh adaptation of the text where these seemed indicated by different customs and racial characteristics, but +, classifications, nomenclature, etc., are strict transla- C tions of the original. / Frederic E. Sondern. or (5 New York City. 24971 TABLE OF CONTENTS PAGE Preface Iv Technic of Clinical Methods for Examination of THE Blood 1 Obtaining the Blood and Preparing the Dried Films 1 Estimation of the Amount of Hemoglobin 2 The Sahli Hemometer 2 Counting the Blood Corpuscles 3 Diluting Fluids 3 Counting the Red Corpuscles 4 Counting the Leucocytes 6 The Differential Count of Leucocytes 7 Examination of Fresh Blood 8 Staining the Dried Films 9 Estimation of the Specific Gravity 10 Formula for Blood Stain 11 Plate I Fig. 1. Development of White and Red Blood Cor- puscles 12 Plate II Fig. 2. Normal Blood 15 Fresh double cover - glass Specimen unstained. Magn. 330 15 Fig. 3. The same specimen. Magn. 750 15 Plate III. — Normal Blood Fig. 4. Stained specimen. Magn. 330 18 Fig. 5. Stained specimen. Magn. 750 18 Plate IV. — The White Corpuscles of the Human Blood. Leucocytes of the Human Blood. Not including Cells of the Bone Marrow 19 Fig. 6. Polymorphonuclear Ncutrophiles. Magn. 750 19 Fig. 7. Lymphocytes. Magn. 750 20 Fig. 8. Transitional Forms. Magn. 750 20 Plate V Fig. 9. Eosinophiles. Magn. 750 22 Fig. 10. Basophilcs. Magn. 750 22 Fitr. n. Turk's Irritation Forms. Magn. 750 23 Iv Plate VI . ^^^^ Fig. 12. Two Eosinophiles and Two Transitionaib. Magii. 750 • • • 24. Fig. 13. Two Transitionals and two Basophiles. Magn. 750 25 Plate VII Fig. 14. Lymphocytes. Magn. 750 25 Fig. 15. Degeneration Forms of Leucocytes. Magn. 750 25 Plate VIII. — The Varieties of Leucocytosis 26 Fig. 16. Neutrophilic Leucocytosis. Moist Double Cover-glass Specimen. Magn. 750 26 Plate IX Fig. 17. Neutrophilic Leucocytosis. Stained Film. Magn. 330 27 Plate X. — Eosinophilic Leucocytosis 28 Fig. 18. Eosinophilic Leucocytosis. Stained Film. Magn. 330 28 Plate XI Fig. 19. Neutrophilic Leucocytosis. Stained Film. Magn. 750 ; • • 29 Fig. 20. Eosinophilic Leucocytosis. Stained Film. Magn. 750 29 Plate XII Fig. 21. Leucocytosis in a Child. Stained Film. Magn. 330 ^^ Plate XIII Fig. 22. Blood Changes in Diphtheria. Stained Film. Magn. 750 32 Fig. 23. Lymphocytosis. Stained Film. Magn. 750. 32 Plate XIV. — The Red Corpuscles of the Human Blood and the Blood Platelets 34 Fig. 24. Erythrocytes. Magn. 750 34 Fig. 25. Nucleated Red Corpuscles (Erythroblasts). Magn. 750 34 Fig. 26. Blood Platelets. Magn. 750 35 Plate XV. — The Anemias Fig. 27. Karyolytic Forms of Erythroblasts. Magn. 750 • • • ^"^ Fig. 28. Polychromatophilic Erythrocytes. Stained Film. Magn. 750 ^'^ V PAGE Plate XVI. — Basophilic Granulations 39 Fig. 29. Basophilic Granulations. Stained Film. Magn. 750 39 Fig. 30. Lead Poisoning. Stained Film. Magn. 750 . 39 Plate XVII. — The Secondary Anemias 41 Acute Form 41 Fig. 31. Acute Anemia. Stained Film. Magn. 750.. 41 Secondary Anemia, Chronic Form 41 Fig. 32. Chronic Anemia. Stained Film. Magn. 750. 41 Plate XVIII Fig. 33. Chronic Anemia. Stained Film. Magn. 330 . 43 Anemia in Children having Congenital Syphilis 43 Fig. 34. Infantile Hereditary Syphilis. Stained Film. Magn. 750 44 Plate XIX. — Secondary Anemias 45 Fig. 35. Chronic Secondary Anemia. Fresh Double Cover-glass Specimen. Magn. 750 45 Fig. 36. Crcnated Erythrocytes. Fresh Double Cover- glass Specimen. Magn. 750 45 Plate XX. — The Primary Anemias 47 Fig. 37. Pernicious Anemia. Fresh Double Cover- glass Specimen. Magn. 750 47 Fig. 38. Chlorosis. Fresh Double Cover-glass Speci- men. Magn. 750 48 Plate XXI Fig. 39. Chlorosis. Stained Film. Magn. 750 49 Fig. 40. Chlorosis. Stained Film. Magn. 750 49 Plate XXII. — Pernicious Anemia 50 Fig. 41. Pernicious Anemia. Stained Film. Magn. 750 51 Plate XXIII. — The Lf.ukf.mias 52 Fig. 42. Myelogenous Leukemia. Fresh Double Cover- glass Specimen. Magn. 750 53 Plate XXIV Fig. 43. Myelogenous Leukemia. Fresh Double Cover- glass Specimen. Magn. 750 54 Fig. 44. Myelogenous Leukemia, Stained Film. Magn. 750 54 Plate XXV. — Chronic Lymphatic I^kukimia 56 Fig. 45. Chronic Lymphatic Leukemia. Stained Fihn. Magn. 330 57 vi Plate XXVI pace Fig. 46. Chronic Lymphatic Leukemia. Fresh Double Cover-glass Specimen. Magn. 750 58 Fig. 47. Chronic Lymphatic Leukemia. Stained Film. Magn. 750 58 Plate XXVII Fig. 48. Chronic Lymphatic Leukemia. Stained Film. Magn. 330 59 Plate XXVIII Fig. 49. Chronic Lymphatic Leukemia. Stained Film. Magn. 750 60 Plate XXIX. — Myelogenous Leukemia 62 Fig. 50. Myelogenous LeuTcemia. Stained Film. Magn. 330 64 Plate XXX Fig. 51. Myelogenous Leukemia. Stained Film. Magn. 750 65 Plate XXXI. — Myelogenous Leukemia 67 Fig. 52. Myelogenous Leukemia. Stained Film. Magn. 750 67 PLATE XXXII. — Myelogenous Leukemia 69 Fig. 53. Myelogenous Leukemia. Stained Film. Magn. 750 70 PLATE XXXIII. — Myelogenous Leukemia 71 Fig. 54. Myelogenous Leukemia. Stained Film. Magn. 750 72 PLATE XXXIV.— Myelogenous Leukemia 73 Fig. 55. Myelogenous Leukemia. Stained Film. Magn. 750 73 PLATE XXXV. — Myelogenous Leukemia 75 Fig. 56. Myelogenous Leukemia. Stained Film. Magn. 750 75 PLATE XXXVL— Acute Leukemia 77 Fig. 57. Acute Leukemia. Stained Film. Magn. 750 . . 78 PLATE XXXVII Fig. 58. Acute Leukemia. Fresh Double Cover-glass Specimen. Magn. 750 80 vii PAGE PLATE XXXVIII. — Blood Changes Associated WITH TUMOES OF THE BoNE MaHROW ■. 81 Carcinoma of the Bone Marrow 81 Fig. 59. Carcinoma of the Bone Marrow. Stained Film. Magn. 330 82 PLATE XXXIX.— Sarcoma of Bone Marrow 84 Fig. 60. Sarcoma of Bone Marrow. Stained Film. Magn. 7.50 85 PLATE XL. — Letikosarcomatosis 86 Fig. 61. Leukosarcomatosis. Stained Film. Magn. 750 87 PLATE XLI. — Blood Parasites. Malarial Parasites and Trypanosomes. Tertian Malaria 88 Fig. 62. Endogenous Development of Tertian Para- site. Magn. 750 .' 88 Fig. 63. Endogenous Development of Quartan Para- site. Magn. 750 ■ ■ ■ 89 Fig. 64. Endogenous Development of Estivo-Autum- nal Parasite. Magn. 750 89 PLATE XLII. — Parasites of Tertian Malaria 91 Fig. 65. Tertian Malaria. Fresh Double Cover-glass Specimen. Magn. 750 91 Fig. 66. Tertian Malaria. Stained Film. Magn. 750. . 92 PLATE XLIII Fig. 67. Tertian Malaria. Stained Specimen. Magn. 7.50 93 Fig. 68. Tertian Malaria. Stained Film. Magn. 750 . . 94 PLATE XLIV Fig. 69. Quartan Malaria. Stained Film. Magn. 750. 95 Fig. 70. Estivo-Autumnal Malaria. Stained Film. Magn. 750 95 PLATE XLV.— Trypanosomiasis 96 Fig. 71. Human Trypanosomiasis. Stained Film. Magn. Approx. 1,000 97 INDEX 98 vni TECHNIC OF CLINICAL METHODS FOR EXAMINATION OF THE BLOOD Obtaining the Blood and Preparing the Dried Films The blood is usually obtained from the lobe of the ear or the tip of the finger by a rapid puncture with one of the various blood lancets or a straight Hage- dorn needle of medium size. Experience develops good technic, and if the procedure is rapid there is practically no pain or subsequent discomfort. Previ- ous cleansing of the skin is desirable, for by thus softening it the puncture is more easily made. The blood should flow slowly of its own accord and with- out pressure on the immediately surrounding tissues. The first drop of blood and any moisture of the skin should be wiped away and the following small drop taken up on the surface of a cover-glass very close to its edge. A slide, wlaich has been carefully cleaned, is held by the left-hand end and the edge of the cover- glass placed in contact with it at the other end, and inchned to the right until the blood drop also touches the shde and spreads the entire width of the cover- glass. Held in this position the cover is now rapidly pushed over the surface of the shde toward the left hand, the drop being drawn along behind it. Any injury to or distortion of the corpuscles is avoided by this method, as no pressure is used, and while the resulting spread is rather thick where begun, it is quite thin and uniform toward the middle and left end of the specimen. Several precautions should be observed. The cover-glass should not come in contact with the skin 1 while it is being charged with blood, and the smaller the amount used, the better the resulting fihn. The cover-glass used for spreading must have an absolutely smooth edge, otherwise the leucocytes may be dis- tributed unevenly, thus jeopardizing the accuracy of the differential count. The whole procedure should be accomplished as quickly as possible so that com- paratively little time elapses between the appearance of the drop of blood and the drying of the prepared fihiis. Estimation of the Amount of Hemoglobin In the apphcation of clinical methods in blood examination, several procedures and appliances for determining the amount of hemoglobin must be con- sidered. There are numerous good methods, but none as yet without some disadvantage. In most of the chnical methods employed, the diluted blood is compared with some artificially colored substance. In the Fleischl-Miescher hemometer this substance consists of a wedge-shaped piece of stained glass, while in the Gowcr's hemoglobinometer a glycerin jelly stained with picro-carmin is used. Both of these methods are trustworthy and universally used, but the new Sahli hemometer deserves special mention. The Sahli Hemometer The inference is reasonable that the best colori- metric determination can be made by comparing the solution to be tested with one of known strength con- taining the identical pigment. While a solution of hemoglobin would be the ideal standard, this is not feasible on account of its rapid deterioration. Con- sequently, Sahli employs a stable hemoglobin deriv- ative as a standard, and converts the blood to be examined into the same derivative before the deter- mination is made. The principle is as follows: One part of blood is mixed with ten parts of decinormal hydrochloric acid in a graduated tube, with the result that an acid heniatin is obtained, quite constant in color and composition. This dark brown fluid dikitcs to a clear yellow with water and is well adapted to colorimetric estimation. The standard solution is sealed in a comparison tube of exactly the same caliber and coiTesponds to a 1 per cent, solution of normal blood. To apply the test, the graduated tube is filled with decinormal hydrochloric acid to the 10 per cent, point, to which 20 c.mm. of blood obtained with the capillary pipette is added, and the whole carefully shaken. As soon as the mixture assumes a clear dark brown color, water is added drop by drop mitil the tint corresponds to that of the standard tube. The percentage of hemoglobin is read from the point of the scale corresponding to the height of the solu- tion. Counting the Blood Corpuscles It is very essential that the blood used for this pur- pose be taken as quickly after the puncture as pos- sible with accurate technic, in order to avoid sources of considerable error. The Thoma-Zeiss counting chamber can be recommended for the separate esti- mation of red cells and leucocytes, but for greater convenience and accuracy in the leucocyte count or when leucocji:es and red cells are to be counted in the same preparation, the chambers ruled according to Zappert or Tiirk are preferable. Diluting Fluids In order to count the red corpuscles it is necessary to dilute the blood with a fluid which will prevent coagulation and cause no change in the cellular ele- ments. One of the following may be used: Hay em's Solution: GRAM Bichloride of Mercury 0.5 Sulphate of Sodium 5.0 Chloride of Sodium 1.0 Distilled Water 200.0 3 Toisson's Solution: Methyl Violet 5 B 0.025 Chloride of Sodium 1.0 Sulphate of Sodium 8.0 Glycerin 30.0 Distilled Water 160.0 For the purpose of counting the leucocytes a dilut- ing fluid is required which lakes the red corpuscles and not only preserves the white ceUs but also makes their nuclei more distinct. This is accomphshed by the use of 0.3 to 0.6 per cent, solution of glacial acetic acid to which a few drops of concentrated aqueous solution of gentian violet may be added to stain the nuclei. Counting the Red Corpuscles The Thoma-Zeiss hemocytometer consists of the following: 1. The Mixing Pipette is a graduated thick-walled capillary tube, pointed at one end and dilated into a bulb at the other, with a short tube at the opposite pole of the bulb to which the rubber aspirating tube is attached. The bulb contains a small glass ball to facilitate the mixing. The capillary tube bears the graduations 0.5 and 1.0 and the upper end of the bulb is marked 101, indicating that the capacity of the bulb is 100 times that of the capillary tube. 2. The Counting Chamber consists of a hea\y glass slide on which a square glass plate, with a circular opening 1 cm. in diameter, is cemented. A circular glass disk 8 mm. in diameter is cemented in the center of this opening in such a way that a ditch separates the two, the slide forming its floor. The surface of the central disk is exactly 0.1 mm. lower than the level of the outer glass plate and forms the floor of the counting chaml)er. Its center is ruled in such a way that 400 small squares are formed each having an area of Hoo sq. mm. An extra line is ruled through each fifth row of squares, as shown in the accompany- 4 ing illustration, to facilitate the counting. The upper limit of the chamber is formed by a superimposed absolutely plane cover- glass. The red cell count is made as follows: After the puncture, made in the usual way, a rather large drop of blood is allowed to col- lect, and this is rapidly - drawn into the pipette to the mark 1.0. Aiter wiping the point, it is plunged into a bottle containing the diluting fluid. Wliile this is being drawn into the bulb the pipette should be twirled between the fingers to mix the blood and diluting fluid. AVhen the mark 101 is reached, aspiration is stopped, the rubber tube re- moved, and with the thumb on the point and the middle finger closing the other end, the whole gently shaken. Some experience is necessary before perfect technic is acquired. As the fluid remaining in the cajjillary tube does not enter into the mixture, the bulb now contains 1 part blood and 99 parts diluting fluid. If the blood has been drawn to the mark 0.5 the dilution is 1 in 200, instead of 1 in 100 as above. The rubber tube is again attached, a few drops blown out, the point wiped dry, and the next small drop placed in the center of the scrupulously clean counting chamber, over which the pohshed cover- glass is placed with some pressure. If the contact is perfect, as it must be if accurate results are desired, the Newton's rings are visible. The drop used must not be too large or some of it will fill the ditch and run between the cover and the outer glass disk. In this case it is necessary to repeat the procedure with a smaller drop. The shde is now left undisturbed for about five 6 minutes to allow the corpuscles to settle on the bottom of the chamber. The counting of the corpuscles is best done with a magnification of about 300, which presents a good picture of the network of squares. The area of each small square is %oo sq. mm., and the distance between the floor and the lower surface of the cover is /4o mm., consequently each square repre- sents wffu cu. mm. The cells in a large number of these squares are now counted, including those which touch or overlap the upper and left-hand boundary lines, but not those which touch or overlap the lower and right-hand boundary hnes. With proper technic the corpuscles are uniformly distributed and counting those in 100 small squares is usually sufficient. If a dilution of 1:100 has been used, the calculation will be as follows: Supposing 1,450 corpuscles were counted in 100 small squares, each square will average Tinr corpuscles. As each square represents a height of /4o mm. and a surface of /4oo sq. mm., each cubic mm. contains 10 X 400 = 4,000 times as many cells. A dilution of 1 :100 having been used the blood con- tains 100 times as many cells as the dilution. There- fore the formula for the calculation will be, 1,450 X 4,000 X 100 = 1,450 X 4,000 = 5,800,000 100 red corpuscles in 1 cu. mm. of blood. The careful cleaning of the pipette is important, and this is accomplished by putting the rubber tube on the capillary tube end of the pipette and drawing water through it. This is followed by alcohol and then by ether, the latter being allowed to run out first one end and then the other by itself. Air is then sucked through })y mouth or asj)irator until the in- terior is dry. The counting chamber is rinsed in water only. Counting the Leucocytes I'^or the enumeration of the leucocytes, large bore pipettes are usually employed, with a resulting dilu- G tion of 1 :10 or 1 :20, the bulb holding but 10 times as much as the capillary tube. After the dilution with the acetic acid solution has been made, the procedure is the same as in counting red corpuscles. For reasons mentioned, a chamber with Zappert ruling should preferably be used for leucocytes, as it admits of counting 2,000 small squares in one speci- men. Supposing the dilution was 1:10, and 2,000 squares were counted, the calculation would be X X 4.000 X 10 ^ X ^ ^„ 2,000 Extreme care and accuracy are absolutely essential in every blood cell count, if the result is to be correct. Beginners cannot possibly avoid all sources of error, and practice is undoubtedly necessary before a proper and uniform technic can be acquired. The Differential Count of Leucocytes Owing to the development in the knowledge of blood changes in disease, a proper estimation of the condition of the blood usually demands information concerning the relative percentage, or the absolute number, of the different varieties of leucocytes pres- ent, in addition to the total leucocyte count. Fluc- tuations in the quantitative relation of the various forms are not uncommon and may present a differ- ential count suggestive or pathognomonic of a given disease. In other cases, valuable prognostic data may be derived from the differential count. Evenly spread and stained blood fihns are em- ployed for the count, and the use of a mechanical stage is desirable. The easiest method of tally is to have another person note the cells as called, but in the absence of such help the number of neutrophiles may be kept in mind and the other varieties noted on paper. With a little practice every one usually de- vises a convenient scheme. A minimum of 300, but preferably 500, cells should be counted in order to 7 secure approximately reliable results. The relative or percentage value of each variety is then figured, and the absolute count should also be determined, by applying the percentages to the total leucocyte count made by means of the counting chamber. ^Vhile making the differential count, the character of the red blood cells, the approximate number of blood platelets, and the presence or absence of nucleated red cells or plasmodia of malaria should also be noted. Examination of Fresh Blood The examination of fresh blood in its moist un- stained state is frequently conducted in an improper manner. When a drop of blood is examined between slide and cover-glass, evaporation and mechanical injury are apt to occur, and the degree of poikilocy- tosis, for example, cannot be properly estimated. Under these conditions the use of an oil-immersion objective is also difficult, as the cover-glass may float on the layer of blood. The following procedure is recommended: A ring of cedar oil is applied around the edge of the concave depression in a hanging drop slide. A perfectly clean square cover-glass is prepared and a likewise clean small piece of cover-glass about 4 mm. in diameter is placed on the center of it. The large bore Thoma- Zeiss pipette is then filled with fresh blood and diluted in proportion of 1:10 with physiological salt solution. After mixing and blowing out a few drops, a little of the diluted blood is allowed to run between the two cover-glasses, as prepared, and the inverted concave slide placed in such a position that the oil ring is in perfect contact with the larger cover-glass. The specimen is then turned right side up and has the appearance as illustrated. The completed specimen of diluted blood is thus prepared in a way which ^certainly prevents evapora- tion and mechanical injury. The capillary layer is 8 preferable to the hanging drop, and the dilution is employed, as it permits a more careful study of the shape of individual corpuscles than is possible when undiluted blood is examined. The specimens may be examined with a low or high power, and if kept warm will show all the characteristics even after the lapse of hours. The excellent results obtained by a-e Counting chamber. Cross section, a. Object slide, b. Counting plate. c. Restangular plates to support cover-glass e.xactly 0.1 mm. above the counting plate, d. Special thick cover-glass. means of this method are well shown in Table II, Figure 3. Staining fluids may be added to the dilut- ing solution for the easier recognition of leucocytes, five drops of an aqueous solution of gentian violet to each 10 c.c. of salt solution being recommended. Staining the Dried Films The Romanowsky method, or one of its numerous modifications, is in almost universal use at present for staining dried blood spreads. For some years the author has used the Leishman* modification almost exclusively. This is an eosinate of methylene blue dissolved in methyl alcohol, which fixes and stains the specimens simultaneously. The Leishman is a panoptic stain, and preferable on account of the simplicity of the procedure. It is used as follows: The air-dried blood fihn, without previous fixation, is covered with about ten drops of staining fluid. After about thirty seconds, double this amount of distilled water is added to and mixed with the stain. Five minutes later the mixture is washed off with water, and the shde dried between hntless blotters. * TV B Leishman. A Simple and Rapid Method of producing Ro- manowsky Staining in Malaria and other Blood Films. British Med. Journ. 9 If the specimen is overstained, the erythrocytes have a deep red or greenish tint. This can be remedied by allowing a few drops of distilled water to remain on the shde for one or two minutes, after which it is washed and dried as before. The entire procedure of spreading the blood, drying, staining and placing the specimen under the microscope should not con- sume more than seven or eight minutes. Only one reagent and a little distilled water are needed for the staining. The results of the method are shown on the following pages, and particular attention is called to Plates IV, V and XV. The stained specimen can now be studied with a low power or with an immersion lens, a permanent mounting in balsam being unnecessary. If balsam is used, it is well to remember that an acid balsam decolorizes the specimen after a time. The cedar oil used on unmounted specimens can be removed with xylol without injury to the blood film. Estimation of the Specific Gravity A knowledge of the concentration of the blood is of considerable importance in a number of diseases. The experienced observer will immediately note de- cided changes in this respect by the appearance of the blood, which observation is sufficiently accurate to determine the degree of dilution best suited to the general blood examination about to be made. The Schmalz method of determining the specific gravity by means of a capillary pycnometer is useful and accurate, but requires experience. A glass tube about 10 cm. long, constricted at both ends and hold- ing 0.2 CO. of fluid, is weighed empty on an accurate balance sensitive to 0.1 mg. It is then filled with distilled water and again weighed. The diff'erence represents the weight of the water held by the tube. After thorough drying the tube is filled with blood and the outer surface carefully cleaned. The net weight of the l)l()od divided by the weight of the water 10 will indicate the specific gravity of the former. The normal specific gravity of the blood is variously stated, but averages 1052 to 1058 in men and 1048 to 1055 in women. Formula for Blood Stain A formula for a polychromatic blood stain was purposely omitted from the ATLAS on account of the difficulty attending its proper manufacture. Pre- pared stains ready for use can be obtained in any of the shops dealing in microscopical goods. Wright's Stain is used in the same manner as that devised by Leishman; the results obtained are the same and the advantage hes in its easier preparation. Dissolve Sodium bicarb., 0.5 gram, in distilled water 100 c.c. and then add Methylene blue rectified 1 gram. Place in a steam steriUzer for one hour. When cold, add about 500 c.c. of a 1:1000 solution of Eosin Griibler w.g., or enough to make the mix- ture purple with a metaUic scum on the surface. The precipitate is collected on a filter and allowed to dry. When thoroughly dry dissolve this precipitate 0.5 gram, in Methyl alcohol Merck reagent 10 J c.c. which makes the stain. Keep balance of the precipitate for subsequent use. 11 PLATE I The Development of White and Red Blood Corpuscles The normal and pathological cells of the human hlood all probably originate in the bone marrow, with the exception of the lymphocytes. Of these a small number are derived from the bone marrow, but by far the larger number come from the lymph glands. A previous study of the mamier of development of the different cells, according to the generally accepted theory, will lead to a better understanding of the chnically important changes in the blood, the signifi- cance of individual cells and the relations they bear to one another. All human blood cells can easily be traced to one parent form ( 1 ) . An unbroken series of cells repre- senting successive stages of development are found between this parent form and the normal cells of the blood. The cells closest to the parent form show different characteristics at an early period by assum- ing either a basophihc (2) or a neutrophihc (3) protoplasm. Subsequently, these cells develop into other forms ( 4 and 5 ) , constituting, on the one hand, a transition into the mature neutrophilic granular cells of the bone marrow (6), the so-called neutro- philic myelocytes, and, on the other hand, pass, by further intermediate stages, into lymphocytes (13) and transitional forms (15). The term "transitional form" is derived from the older interpretation of this cell as a preliminary stage of the polymorphonuclear neutrophile, or as an intermediate stage between that cell and the neutrophilic myelocyte. This view is 12 Plate I. Fig. 1 \i I't scarcely justified at the present state of our knowl- edge and it is probable that the transitional form also represents a terminal cellular type incapable of fur- ther development. The large cell with homogeneous basophihc proto- plasm (2) also gives rise, by way of intermediate stages, to the basophihc myelocj'te (8), which in turn is transformed into the basophile (11). The eosino- phile (12) may be derived, by way of the eosinophilic myelocyte (9), from the prehminary stages of the basophihc myelocji;e. A close relationship probably exists between the basophihc and the eosinophihc myelocyte, notwithstanding their dissimilar staining quahties, as intermediate stages between these two cells are not infrequently encoimtered in leukemic blood. Basophiles are invariably found increased in cases of relative eosinophiha, and the association of these mature cells is a common chnical phenomenon. The neutrophihc leucocyte (10) is derived from the neutrophihc myelocji;e (6) in the same way as the normal eosinophil and basophile (12 and 11) develop from the corresponding eosinophihc myelocyte (9) and the basophihc myelocyte ( 8) . The rare irritation form of Tiirk (14) is at present interpreted as a lymphocyte, the further development of which is due to a pathological stimulus, and not as a prehminary stage of the mature lymphocyte (13), as formerly beheved. The erythrocytes can also be traced, by direct transition, to the parent cell; the large mononuclear cell with homogeneous basophihc protoplasm (2) being a forerunner of the megaloblast (16). The megaloblast in its early stage may have a voluminous basophihc cell body and a relatively large nucleus and, occurring in the peripheral circulation, is often distinguished with difficulty from the Tiirk irritation form (see Fig. 41). As the result of the further development of the megaloblast, the normoblast is formed, and subsequently the erythrocyte (17). 13 Plate I represents a genealogical scheme of devel- opment, including the chief types only. In the desig- nation of cells the suggestion of Grawitz is recom- mended, that for the present the terms having definite significance, such as "myelocyte," be adhered to, with the additional characteristic attached, for example: neutrophihc myelocyte, eosinophilic myelocyte, etc., rather than the arbitrary nomenclature suggested by different authors. In the development of all blood corpuscles, the following chief forms must be considered: 1. Large mononuclear cell with pale homogeneous cell body = parent cell. 2. Large mononuclear basophihc cell. 3. Large mononuclear neutrophilic cell. 4. Mononuclear basophihc cell in which there are few neutrophihc granules. 5. Mononuclear neutrophihc cell in which there are few neutrophihc granules. 6. Mononuclear cell with dense neutrophihc gran- ulation = neutrojjhihc myelocyte. 7. Mononuclear basophihc cell with beginning basophihc granulation. 8. Mononuclear cell with basophilic granulation = basophihc myelocyte. 9. Mononuclear cell with eosinopliihc granula- tion = eosinophilic myelocyte. 10. Neutrophile. 11. Basophile. 12. Eosinophile. 13. Lymphocyte. 14. Tiirk irritation form. 15. Transitional form. IG. Erythroblast. 17. Erythrocyte. 14 Plate II. Figs. 2, 3 PLATE II NORMAL 15L00D The blood of healthy individuals presents certain physiological variations within comparatively narrow limits. The average composition is approximately as follows : Children >4 to 15 years: Hemoglobin, 70 to 80 per cent.; red cells, 4,900,000; leucocytes, 9,000 in 1 c. mm. Men: Hemoglobin, 90 to 100 per cent.; red cells, 5,100,000; leucocytes, 7,500 in 1 c. mm. Women: Hemoglobin, 85 to 95 per cent.; red cells, 4,500,000 to 5,000,000; leucocytes, 7,500 in 1 c. mm. Healthy Man, 32 years old: Hemoglobin, 120 per cent.; red cells. 5,600,000; leucocytes, 7,800. Figure 2.— Fresh Double Cover-Glass Specimen Unstained. Magnification 330 The pale yellow color of the erythrocytes is the characteristic normal hemoglobin tint. The cells are uniform in size, some appearing cup shaped on ac- count of their position. The detailed structure is more apparent when a higher power is used. Figure 3.— The Same Specimen. Magnification 750 Normal red blood corpuscles appear cup-shaped with thickened walls. This appearance is only noted as long as injurious influences are excluded, such as evaporation, drying from any cause, hypertonia, isotonia, cold or heat, which cannot always be pre- vented, even by experienced workers. This cup shape 15 is best noted in the corpuscles lying by themselves; the convexity is apparent, and on the opposite side the concavity of the cup can be seen, its rounded margin merging into the outer wall of the blood cell. The lateral aspect of the red corpuscle is well curved and sausage shaped, on the concave side of which there is a hght transparent wall with convex margin. If the corpuscle hes so that the open end of the cup can be looked into, the appearance is that of a circle with a sharply defined, lighter colored center. This light center corresponds to the trans- parent crest at the floor of the cup, and the darker colored outer ring to the relatively much thicker wall of the cup. Seen from above the corpuscle appears globular, and the cup -shaped form can only be noted on focusing a lower level. Most of the red corpuscles in the double cover-glass specimen present this appearance. Deviations from this fundamental form are noted, but these have as yet assumed no pathological sig- nificance. The cup shape may be more or less pro- nounced and the opening may vary in width. The erythrocyte is frequently seen in the form of a biconcave disk, both concavities occupied by a con- vex transparent membrane. This appearance is ex- plained by the crest of the bell having become in- dented by contact with another corpuscle. These indentations are rarely seen in the lateral wall of the corpuscle. AVlien the red corpuscles are close to- gether the so-called rouleaux formation occurs as the result of crowding, the concave portion of one cor- puscle slipping over the convex portion of the adja- cent one. In this condition the actual form of the corpuscles cannot be made out. (See Figs. 16, 35, 37, 38, 42, 43, 46 and 51.) {Weidenreich.) Three leucocytes are also seen in the field; the smallest, with a round nucleus surrounded by a nar- row rim of i^rotoplasm, is a lymphocji;e. The larger 16 cell on the left with polymorphonuclear configuration and dense finely granular protoplasm is a jjoly- morphonuclear leucocyte. The cell above presents a striking appearance, the nucleus consisting of several portions connected by thin nuclear strands. The protoplasm is filled with coarse, round, strongly refractive granules, some lying apparently beyond the cell but actually within protoplasmic processes of the cell, which is in slight ameboid motion. This cell is an eosinophile which is identified in the fresh specimen by its highly refractive coarse granulation. 17 PLATE III NORMAL BLOOD Figure 4. — Stained Specimen. Magnification 330 [The same case.] All the erythrocytes are uniform in size and, on account of being spread, are seen as flat circular yellowish-red disks, the cup shape not being apparent. While some seem uniformly pigmented, the majority have a lighter colored central zone and a darker periphery, which is due to the original cup shape of the corpuscle. The details are more apparent with the use of a higher power. Figure 5. — Stained Specimen. Magnnification 750 [The same case.] The appearance of the erythrocytes is more dis- tinct in this specimen. The depressed center, being a thin wall, is faintly stained, while the thicker edge has a deeper tone. A neutrophilic leucocyte and two lymphocytes of different size are also seen in the field. 18 Plate III. Figs. 4, 5 Plate IV. Figs. 6, 7, 8 PLATE IV THE WHITE CORPUSCLES OF THE HUMAN BLOOD Leucocytes of the Human Blood Not Including Cells of the Bone-Marrow The number of leucocytes in 1 c. mm. of blood in an adult is approximately 7,500. These consist of a variety of different forms, which, under normal cir- cumstances, are present in a definite numerical ratio to one another. This enumeration is known as the "Differential Count of Leucocytes," and the follow- ing are the normal varieties and figures : Polymorphonuclear Neu- trophiles 65 to 70 per cent, or 4,900 to 5,300 in 1 c.mni. Lymphocytes 20 to 25 per cent, or 1.500 to 2,000 in 1 c.mni. Transitional Forms 3 to 5 per cent, or 230 to 380 in 1 c.min. Eosinophiles 2 to 4 per cent, or 150 to 300 in 1 c.mm. Basophiles % per cent, or 40 in 1 c.mm. Turk's Irritation Forms rarely occur. In children under 5 years of age, the lymphocyte is the predominating cell, and the polymorphonuclear neutrophiles usually amount to less than 50 per cent. Figure 6. — Polymorphonuclear Neutrophiles, Magnification 750 These cells are usually about twice the size of an erythrocyte, rarely larger. The nucleus is character- ized by its polymorphous configuration, some of the portions often being connected by thin nuclear strands. Apparently separate nuclear segments are occasionally seen, but these have "underground" connection, as the cells are never polynuclear. The 19 protoplasm is neutrophilic and shows close fine granulations which are especially apparent in cases of leucocytosis, but are occasionally so shght that they' cannot be brought out even in overstained specimens. The neutrophihc quaUt^- of the proto- plasm also varies, and these differences are probably due to different ages and functions of the cells, rather than to any fault in staining. Figvire 7. — Lymphocytes. Magnification 750 The cells are extremely varied in size and some show a narrow, others a broad ring of protoplasm. The majority are small, little larger than an eryth- rocyte, but very large forms, four or five times the size of the small ones, are frequently seen in children, and occasionally in adults. The nucleus is usually round and sometimes indented, and in the larger forms it is oval or polygonal. It stains deeply in the small cells and rather faintly in the larger ones and two or more nucleoli may become visible. The protoplasm is shghtly basophihc, — more so in the small cells than in the large ones, — and occasionally the outer zone stains darker than that close to the nucleus. The large Ijinphocytes may present angular or irregular outlines. A smaller or larger nimiber of all lympho- cytes show acidophilic granulation, which appears as fine particles or angular granules. This is noted in every specimen and has no pathological significance as yet. Figure 8. — ^Transitional Forms. Magnification 750 These cells are invariably larger than the poly- morphonuclear neutrophiles and frequently irregu- larly round in outline. (The irregular outline as in the case of the large lymphocytes is probably referable to adjacent erythrocytes, or those in close proximity.) 20 Compared with the other lymphocytes, the nucleus is always paler and poorer in chromatin. It is usually horseshoe-shaped or multilobular, rarely round and indented. The protoplasm is slightly basophilic and contains more or less marked neutrophilic granulation most apparent in the nuclear indentations. With these characteristics in mind, the cells are easily dif- ferentiated from the polymorphonuclear neutrophiles. 21 PLATE V Figure 9. — Eosinophiles. Magnification 750 These are very striking and easily identified round cells of the same size as polymorphonuclear neutro- philes, rarely larger or smaller. They frequently have two or three oblong or oval nuclear portions con- nected by thin nuclear strands, which are occasionally unequal in size. The protoplasm is faintly basophihc and shows a bright-red coarse granulation. The granules are round, varying in number, and may be arranged in groups. Some cells are so densely filled with granules that they can scarcely be distinguished. In one case of trichinosis, the author found a cell containing an equal number of eosinophilic and basophilic granules (see Fig. 9). Figure 10. — Basophiles. Magnification 750 These cells vary in appearance and are about the size of an eosinophile. They occur less frequently, and are not as easily recognized. The nucleus occupies about two-thirds of the cell, usually pre- senting the outline of a clover-leaf or rosette, rarely round or indented, with no defined margin. The protoplasm shows a fine basophilic or eosinophilic network. Round coarse granules of varied size are seen at the points of intersection, or these may fill the entire cell and lie on the nucleus. The granules take a deep chromatin stain and have basic properties. The cells are occasionally free from granules and then vacuoles are found in the network. 22 I- Plate V. Figs. 9, 10, 11 ■■^.: n # ^ &> Figure 11.— Tiirk's Irritation Forms. Magnification 750 These cells are only found in pathological blood after leucocytoses and inflammatory processes. They resemble the large lymphocytes in type, have a large nucleus, poor in chromatin and frequently small nucleoli. The nucleus is not always in the center of the cell and is surrounded by a band of protoplasm which varies in width and takes a dark blue stain. Vacuoles may be found in the cell body. Amitotic nuclear division figures are not uncommon. 23 PLATE VI Figures 12 to 14 show some of the above described leucocytes, as well as a comparison in appearance and size between one another, and the erythrocytes. Figure 12. — Two Eosinophiles and Two Transi- tionals. Magnification 750 In the eosinophiles, the characteristic configuration of the nucleus and the arrangement of the distinctly stained granules are particularly evident. The two transitionals present characteristics by means of which they are easily distinguished from polymor- phonuclear neutrophiles (see Fig. 6). A few blood platelets are seen on the left. Figure 13. — Two Transitionals and Two Basophiles. Magnification 750 The upper transitional appears as a large mono- nuclear leucocyte. Close inspection shows an in- dented and constricted nucleus. The lower transi- tional has a lobular nucleus and is probably a more mature cell. Two basophiles are seen on the right with differ- ently shaped nuclei and coarsely granular protoplasm. 24 Plate VI. Figs. 12, 13 Plate VII. Figs. 14, 15 PLATE VII Figure 14. — Lymphocytes. Magnification 750 The lymphocytes shown in the picture represent an infrequently observed type, and may occasion diagnostic difficulty. The two upper cells illustrate the maximum size of the usually much smaller lym- phocyte. The nuclei are poor in chromatin and show distinct nucleoli. The acidophilic granules are well brought out by the Romanowsky stain in all three of the cells. Figure 15. — Degeneration Forms of Leucocytes. Magnification 750 Two neutrophiles are seen above, two lymphocytes to the left, a transitional below, a Turk's irritation form on the right and an eosinophile in the center. All these cells show vacuoles varying in size and number. These changes are usually noted in speci- mens from much debilitated or moribund patients many hours before death. One of the polymorpho- nuclear neutrophiles in the picture has been dis- tended, and finally ruptured by the increase in size of the vacuoles. In the transitional, the zonal stain and the marginal arrangement of the vacuoles are noteworthy, and probably due to lessened resistance at the periphery of the protoplasm. The occurrence of this vacuolar degeneration in a large number of leucocytes would thus seem to be a bad prognostic sign. 25 PLATE VIII THE VARIETIES OF LEUCOCYTOSIS Neutrophilic JLeucocytosis The diagnosis of this condition is based not only on an increase in the relative percentage of polymor- phonuclear neutrophils in the differential count but also on an absolute increase as well. The lowest figures which constitute a neutrophihc leucocytosis are approximately 8,000 neutrophiles in 1 c.mm., the total leucocyte count being about 10,000. The maxi- mum figures are indefinite, but counts over 60,000 neutrophiles in 1 c.mm. are rare. This is the most common type of leucocytosis. The lymphocytes and eosinophiles are usually decreased both in relative percentage and in absolute numbers. Female, 16 years old. Pleurisy and Exudative Pericarditis. I,eucocytes, .34,200 in 1 c.mm. Figure 16. — Neutrophilic Leucocytosis. Moist Double Cover-glass Specimen. Magnifica- tion 750 The erythrocytes are normal in appearance. The white corpuscles, relatively increased in number as con)j)ared to tlie erythrocytes, arc seen to be large polymorphonuclear cells with finely granulated proto- plasm, indicating that they are polymorphonuclear neutrophiles. Some show ameboid motion. 26 Plate VIII. Fig. 16 Plate IX. Fig. 17 PLATE IX Figure 17. — Neutrophilic Leucocytosis. Stained Film. Magnification 330 [The same case.] The red corpuscles show slight variations in color due to the shght anemia. Sixteen polymorphonuclear neutrophiles and one transitional are seen in the field. The hlood platelets are somewhat increased in number. 27 PLATE X EOSINOPHILIC LEUCOCYTOSIS The increase in the number of eosinophiles in this type of leucocytosis is relatively less marked than the increase of neutrophils in neutrophilic leucocytosis, and corresponds approximately to the relative pro- portions in normal blood in which there are about 150 to 300 eosinophiles in 1 c.mm. The presence of 6 per cent, or more eosinophiles in the differential count constitutes an eosinophiha when the absolute count is correspondingly increased. This condition is noted chiefly in bronchial asthma, skin disease, helminthiasis and particularly in trichinosis. Male, 31 years old. Trichinosis. Total leucocyte count 18,000 in c.mm. Differential count shows 49.6 per cent, eosinophiles or 8,930 in 1 c.mm. Figure 18. — Eosinophilic Leucocytosis. Stained Film. Magnification 330 Erythrocytes unchanged. Numerous leucocytes are seen in the field, the majority (17) of which are eosinophiles, which are easily identified though the granular stain is imperfect. Four neutrophiles are seen in the center and above these there are two lymphocytes and one transitional. 28 PLATE XI Figure 19. — Neutrophilic Leucocytosis. Stained Film. Magnification 750 The polymorphonuclear type of the cells indicates that they are neutrophiles. The granulation of the protoplasm is denser than in normal blood, a change usuallj'^ noted in leucocytosis. The blood platelets vary in size and are increased in number. Figure 20. — Eosinophilic Leucocytosis. Stained Film. Magnification 750 [Till' same case as Figure 18.] The decided increase in the number of eosinophiles is evident. The cells are somewhat smaller than the usual eosinophile, and some shovp^ granulations on the nucleus. 29 PLATE XII Girl, 3 years old. Chronic intestinal catarrh. Leucocytes 14,000 in 1 c.mm. Diflferential count shows 48 per cent, lym- phocytes or 6,500 in 1 c.mm. Figure 21. — Leucocjrtosis in a Child. Stained Film. Magnification 330 While a true neutrophilic leucocytosis may occur in children, the simultaneous increase in neutrophiles a. I'olyniorphonuclear Neutrophile*. 6. Lympliocytes. o. Transitionals. d. Tiirk'a irritation forma. 80 Plate X. Fig. 18 Plate XI. Figs. 19, 20 and in mononuclear forms, particularly the lympho- cytes, is the most freciuent type of leucocytosis. Atypical forms of leucocytes are frequently seen in these cases, such as neutrophiles with round nuclei, Tiirk's irritation forms, and myelocytes, their number serving as an index to the gravity of the disease. The red corpuscles show no particular change. 31 PLATE XIII Child, 6 years old. Septic diphtheria of nose, fauces and larynx. Duration 8 days. Cutaneous hemorrhages. Blood ex- amination 6 hours before death shows: Hemoglobin, 100 per cent.; red cells, 4,388,000; leucocytes, 72,000. Figure 22. — Blood Changes in Diphtheria. Stained Film. Magnification 750 In fatal cases of diphtheria in addition to the usual neutrophilic leucocytosis, numerous neutrophilic mye- locytes are found, while these are not noted in favor- able cases. Turk's irritation forms and mononuclear neutrophiles also may occur in the favorable cases, and have no particular significance, but the presence of myelocytes must always be regarded as an unfavor- able prognostic sign. Six neutrophilic leucocytes are seen in the field; to the right there is a small lymphocyte with scarcely visible protoplasm, toward the center a distorted transitional, and adjoining this there are two neu- trophilic myelocytes, their irregular outline due to pressure. The red corpuscles show evidences of some anemia. Male, 22 years old. Typhoid fever, 26th day. Hemoglobin, 80 per cent.; red cells, 4,510,000; leucocytes, 6,300. Differential Count: Neutrophiles, 38.5 per cent, or 2,400; lymphocytes, 53.8 per cent, or 3,400; transitionals, 5 per cent.; eosinophiles, 2.4 per cent. ; basophiles, none ; irritation forms, 0.3 per cent. Figure 23. — Lymphocytosis. Stained Film. Magnification 750 Lymphocytosis consists of an increase in the rela- tive percentage as well as in the actual number of 32 Plate XII. Fig. 21 normal lymphocytes, which are derived in large num- bers from lymph glands by virtue of pathological change or stimulus. The condition is observed in whooping cough, rickets, syphihs and typhoid con- valescence, and follows injections of tubercuhn. In the illustration, the red cells show pale centers owing to diminished hemoglobin. Three lympho- cytes, and one neutrophile practically without gran- ules, are seen in the field. The lymphocytes all belong- to the large type, and one shows shght acidophihc granulation of its protoplasm. 33 PLATE XIV THE RED CORPUSCLES OF THE HUMAN BLOOD AND THE BLOOD PLATELETS Figure 24. — Erythrocytes. Magnification 750 Normal Erythrocytes are round disks with an aver- age diameter of 7 to 7^ microns, the margin often staining deeper than the center. Microcytes are abnormally small ery1;hrocji;es, round or deformed, with a normal or pale hemoglobin tint. They are to be considered degeneration forms. Macrocytes are abnormally large erythrocytes, usually quite round, frequently polychromatophilic, and rarely showing a uniform hemoglobin tint. They represent preliminary stages of the normal erythrocyte. Poikilocytes are deformed erythrocytes, usually bottle or pear shaped, and often very irregular. The hemoglobin content is always more or less diminished. They are considered degeneration forms which have lost their normal contour on account of lessened resistance. Figure 25. — Nucleated Red Corpuscles (Erythro- blasts). Magnification 750 Erythroblasts represent preliminary stages of nor- mal red blood cells, their appearance in the periph- eral blood indicating increased activity of the bone marrow in the formation of erythrocytes. Normoblasts are of the same size as erythrocytes and have a relatively large round nucleus showing 84 Plate XIII. Fig.. 22, 23 radiating or segmented structure. The cell body of the mature normoblast has a pure hemoglobin tint, but the younger cells are frequently polychromato- philic. 3Iicroblasts are characterized by their small size, and show degenerative changes of the cell body (poikilocytosis), and pycnotic nuclei of dense struc- ture, which stain deeply. Free microblast nuclei are sometimes seen, and are easily differentiated from other structures by their small size and dark blue color. Megalohlasts are two or three times the size of normoblasts, and the older forms have a relatively small nucleus. Polycliromatophihc megalohlasts are not uncommon, and present large, loose structured nuclei, poor in chromatin. The megaloblast origi- nates from a pathologically altered regeneration of the blood with a reversion to the embryonic type. Nuclear Division Figures of Erythroblasts. Ami- totic (direct) division is more frequently observed in megalohlasts than in normoblasts. Attention to the hemoglobin or polychromatophihc stain of the cell body differentiates these cells from lymphocji;es of similar appearance, in which the nucleus may also undergo amitotic division, but the cell body is always basophihc (blue). Mitotic division is rarely observed in the leukemias and in pernicious anemia. The nuclear division figures are often large and beautifully marked with more or less distinct diaster formation. The cell body is still slightly basophilic and usually shows basophilic granulation. Figure 26. — Blood Platelets. Magnification 750 These are blood elements of various shapes, which range in size from some scarcely visible, to others with a diameter of 10 microns. The smallest forms appear as pale, sharply circumscribed disks, which 3S may have crenated margins, and show a nuclear sub- stance also varying in size and often with only few chromatin granules. The larger forms are oval or rod-shaped, with abundant nuclear substance of a dark chromatin tint. These may show processes varying in length up to twice the diameter of the platelet, which also contain chromatin (see Fig. 40). Ordinarily, the platelets occur in small groups, or singly, but in the blood of persons having mahgnant tumors they may be found aggregated in large numbers. The origin and the significance of the platelets are still obscure. They are usually foimd increased in number during, and after, all conditions associated with leucocytosis, or with disintegration of leucocytes. They are enormously increased in myelogenous leukemia. The practical method of determining their number is by means of an approximate estimate in the stained fihn*. 86 Plate XIV. Figs. 24, 25, 26 26 ■*1 V '«3 <• «»> ^ « a , 'i- 4 1 \ ••?:t^^_; ' ■ • " ■ PLATE XV THE ANEMIAS Polychro matopMUs This term indicates a changed behavior of the erythi-ocytes toward the stain. PolychromatophiUc erythrocytes show a mixed staining quahty varying from an ahnost pm-e eosin to a dirty blue or violet. In the more marked instances the corpuscles appear spotted as if occupied by a basophilic stroma. The condition is found in all stages of development of the immature erythrocyte, particularly in the nucleated cells, and those showing nuclear division figures. It is, therefore, regarded as a sign of imma- turity of the red corpuscle, and consequently a symp- tom of increased regeneration. Figure 27. — Karyolytic Forms of Erythroblasts. Magnification 750 These are often present in large numbers in cases of progressive severe anemia, and particularly in the anemia of congenital syphiMs in children. The nuclei are extremely varied in appearance and show budding and constrictions with pycnotic seg- ments, A small nucleus, poor in chromatin, is some- times seen in a polychromatophiHc cell, apparently a remnant of an original nucleus which has undergone dissolution. v Cliild, 2J/2 years old. Anemia gravis with congenital syphilis. Red cells, 1,564,000; leucocytes, 14,000. Figure 28. — PolychromatophiHc Erythrocytes. Stained Film. Magnification 750 The erythrocytes show moderate variations in size and shape, with both slight and pronounced poly- 37 chromatophilia (see Fig. 41). A megaloblast in process of division, with polyclu'omatophilic cell body, is seen above, and on the right there is a lymph- ocyte with the usual blue stained protoplasm. 88 Plate XV. Figs. 27, 28 Plate XVI. Figs. 29, 30 PLATE XVI BASOPHILIC GRANULATION In this change the normal erythrocyte, or more frequently the polychromatophilic erythrocyte, con- tains granules whicli take a basic stain. The granules vary in size; some are scarcely visible and others are as large as the neutrophihc granules in leucocytes. Poikilocytes and microcytes occasionally contain unusually large angular granules. They also vary in number, sometimes filhng the entire cell. Basophihc granulation is also found in nucleated red cells and in those presenting nuclear division figures. Polychromatophiha being considered an indication of immaturity, the designation of an accompanying basophilic granulation as "granular degeneration" does not seem justified, but as yet no definite opinion is possible. It is observed in all types of anemia and particularly in cases of lead poisoning. Figure 29. — Basophilic Granulation. Stained Film. Magnification 750 Composite picture of specimens from a case of chronic lead poisoning. The varieties of basophilic granulation and its combination with polychroma- tophiha are the noteworthy features. Male, 25 years old. Painter, suffering from chronic lead poisoning. Figure 30. — Lead Poisoning. Stained Film. Magnification 750 Granular erythrocytes are invariably present in cases of lead poisoning. They are more numerous in 39 this condition than in any other disease, and the blood picture is practically pathognomonic. The erythrocytes show slight variations in size and color. In some the color is limited to the margins and shght polychromatophilia is apparent. Nine red cells which have more or less marked basophilic granulation are seen in the field. 40 PLATE XVII THE SECONDARY ANEMIAS Secondary anemia includes all the anemic condi- tions developing in the course of different diseases which are not based on the primary involvement of the blood-making organs. The gi'oup includes all affections leading to loss of blood, mahiutrition and cachexia (gastric and intestinal ulcers, animal para- sites, infectious diseases, malignant tumors). In order to study the morphology of the red corpuscles, in primary as well as secondary anemia, it is essential to examine fresh specimens, as the changes noted in stained films are often artefacts. Secondary Anemia. Acute Form Soon after a severe hemorrhage there is a diminu- tion in the number of red corpuscles and in the amount of hemoglobin, due to a transfer of fluids from the tissues into the blood. Regenerative phe- nomena are soon noted by the presence of polychro- matophilia, macrocytes and normoblasts, which im- mature forms are usually deficient in hemoglobin. The number of red cells increases rapidly, while the increase in hemoglobin content is slower. Female, 45 years old. Ulcer of the stomach. Hemoglobin, S.'j per cent.; red corpuscles, 1,960,000; leucocytes, 6,300 in 1 cram. Figure 31. — Acute Anemia. Stained Film. Magnification 750 The erythrocytes are further apart in the stained specimen, owing to the dilution of the blood. The 41 lack of uniformity in size is due to the presence of numerous polychromatophilic macrocytes, and not to changes in the normal cells. A polychromato- philic normoblast is seen above and a lymphocyte be- low. Several blood platelets are also present. Secondary Anemia. Chronic Form In addition to the changes noted above, these cases also show variations in the shape of the red cells and a more or less pronounced diminution in the amount of hemoglobin. The red cells may simply show decided loss of hemoglobin, without polychro- matophiha. An absence of immature erythrocytes in cases of secondary anemia justifies a suspicion of paralysis of function of the bone marrow, as far as formation of erythrocytes is concerned. The more numerous the immature red cells, the greater the functional activity of the blood-forming organs. When the bone marrow is suddenly called upon to supply a marked deficit, there may be a transitory flooding of the circulation with many nucleated cells. Male, 54 years old. Carcinoma of stomach. Hemoglobin, 35 per cent.; red cells, 1,336,000; leucocytes, 13,100 in 1 c.mm. Figure 32. — Chronic Anemia. Stained Film. Magnification 750 The red cells show the characteristic evidences of secondary anemia, i. e., diminished and uneven hemoglobin distribution, polychromatophilia, changes in size and shape of corpuscles. 42 Plate XVII. Figs. 31, 32 Plate XVIII. Figs. 33, 34 PLATE XVIII Figure 33. — Chronic Anemia. Stained Film. Magnification 330 The same changes are noted as in Fig. 32, and the pathological condition is particularly apparent when compared with Fig. 3, which represents normal blood. Two neutrophils are seen in the field, with a normoblast above and numerous blood platelets. Anemia in Children having Congenital Syphilis In well marked cases the changes in the red cells are sufficiently pronounced to make this one of the cardinal symptoms. In addition to a leucocytosis, numerous bone-marrow cells are found, which justify the l)ehef that the bones are also involved in the pathological lesion and that the presence of these cells is the result of direct irritation of the bone marrow by the local syphilitic process. Neutro- philic and eosinophilic myelocytes and immature lymphocytes are present, together with a large num- ber of nucleated red cells, microblasts, normoblasts and frequently megaloblasts and nuclear division figures. The large number of karyolytic forms is noteworthy (see Fig. 27). The red cells appear de- ficient in pigment, but poikiloc_\i;osis, polychromato- philia, and basophilic granulation are not common. The number of erythrocytes is always considerably diminished, with a corresponding reduction in the amount of hemoglobin. 43 Child, lyz years old. Hereditary syptilis. Red cells, 2,500,000; leucocytes, 18,000 in 1 c.mm. Figure 34.— Infantile Hereditary Syphilis. Stained Film. Magnification 750 The specimen contained numerous nucleated red cells their number exceeding the total leucocyte count. The stained film of blood, obtained after death, shows some disintegrating cells. Normoblasts and Megaloblasts. Some of the latter unusually large, with more or less basophilic protoplasm, and one or more nuclei poor in chromatin. Karyolytic Forms of Normoblasts and Megaloblasts. Lympliocytes. One with diviiicd Nucleus. Granular and Non-granular Bone marrow Cells. PLATE XIX SECONDARY ANEMIAS Figure 35. — Chronic Secondary Anemia. Fresh Double Cover-glass Specimen. Magnifica- tion 750 [Same case as Figure 32.] The pale tint of the red cells indicates their loss of hemoglobin, especially when compared with nor- mal blood. The varied outline of red corpuscles is most appar- ent in the fresh specimen, but an opinion as to their size requires experience. Pear-shaped and bottle- shaped cells, and multilobular poikilocytes are pres- ent, A microcyte is seen in the center of the field. The two humped and oblong red cells are changes, due presumably to evaporation, the details of which are seen in the following picture. Figure 36. — Crenated Erythrocytes. Fresh Double Cover-glass Specimen. Magnification 750 The illustration shows the crenated forms of red cells, formerly believed to be the same thing as poikilocytes. The characteristic changes are evident. The crenated forms may be observed in any specimen of fresh blood which has evaporated or suffered mechanical injury. In fact the occurrence of a few of these near the margin can scarcely be prevented in the fresh specimens, though carefully prepared. Crenation within a few seconds of all the red corpuscles in a properly made specimen is pathologi- 45 cal, and must be considered an evidence of lessened resistance on the part of the erythrocytes. Error is prevented by observing this in several successively prepared specimens. Normal blood does not show this change to any extent until after the lapse of from thirty minutes to several hours. 46 Plate XIX. Figs. 35, 36 Plate XX. Figs. 37, 38 PLATE XX THE PRIMARY ANEMIAS The characteristic feature in the diagnosis of primary anemia, is the presence of anemic conditions, varying in kind and degree with an absence of all other causative diseases. In chlorosis and certain anemias of childliood, nothing but an affection of the blood itself has as yet been demonstrated, whereas in pernicious anemia the blood-forming organs are the seat of the lesion. The anemias of childhood present no specific or particularly characteristic changes in the blood, but in chlorosis, as well as in pernicious anemia, definite evidences are found, which are characteristic of the one or the other condition. Female, 63 years old. Pernicious Anemia. Hemoglobin, 35 per cent.; red cells, 1,032,000; leucocytes, 16,000 in 1 c.mm. Marked polychromatophilia and pronounced variations in the size of the red cells. Normoblasts and megaloblasts are present. Figure 37. — Pernicious Anemia. Fresh Double Cover-glass Specimen. Magnification 750 Most of the red cells are deficient in hemoglobin, but some have a normal color. The uneven dis- tribution of hemoglobin is always apparent. The pronounced difference in the size of the corpuscles is noteworthy ; about half of them are megalocytes, and there are also many microcytes, some so small that they are difficult to distinguish from blood platelets in the fresh specimen. Poikilocytosis was not a pronounced feature in this 47 case of pernicious anemia in which the diagnosis was verified by autopsy. This change is evidently not present to the same degree in every case, and probably depends to some extent on the duration of the disease. Female, 28 years old. Chlorosis. Hemoglobin, 50 per cent.; red cells, 4,320,000; leucocytes, 9,600 in 1 c.mm. Figure 38. — Chlorosis. Fresh Double Cover-glass Specimen. Magnification 750 In chlorosis, even the fresh blood differs in several essential features from that of secondary anemia or of pernicious anemia. Primarily, the erythrocytes show a uniform decided diminution in blood-coloring mat- ter, which is very apparent in severe cases. Poildlo- cji;osis is not as pronounced in severe chlorosis as it is in severe secondary anemia, and the marked difference in the size of the corpuscles seen in perni- cious anemia, is absent. In cases of severe hydremia a uniform increase in the size of a large number of erythrocytes is frequently observed. ^^Hien the chlor- osis is not jjronounced, no particularly characteristic changes can be made out in a fresh specimen. Figure 38 shows a specimen of blood from a case of chlorosis. The red cells are particularly uniform in size and very pale in color. Few pear-shaped poiki- locj'tes are present. The erythrocji:es, viewed later- ally, show a hght zone which is probably a reflection. 48 Plate XXI. Figs. 39, 40 PLATE XXI Girl, 26 years old. Severe Chlorosis. Hemoglobin, 35 per cent.; red cells, 4,000,000; lymphocytes, 4,400 in 1 c.mm. Figure 39. — Chlorosis. Stained Film. Magnification 750 In chlorosis the diminution in the amount of hemoglobin is much greater than in the number of red corpuscles, and this characteristic loss of coloring matter is apparent in all the corpuscles by the narrow stained rim as shown in the illustration. There is only a sHght difference in the size of the cells, but poi'kilocytosis is evident. The platelets are increased in number. A lymphocyte is seen on the right. Female, 26 years old. Convalescing Chlorosis. Hemoglobin, 90 per cent. ; red cells, 4,600,000 ; leucocytes, 7,200 in 1 c.mm. Figure 40. — Chlorosis. Stained Film. Magnification 750 The specimen shows the decided and uniform in- crease in the amount of hemoglobin in the corpuscles during convalescence, but the pale centers indicate that this is not yet quite normal. Few poikilocytes are present. The flagella-like processes of the platelets are seen in this specimen. 49 PLATE XXII PERNICIOUS ANEMIA In cases of pernicious anemia the blood as a rule presents a characteristic series of changes not noted in any other disease. The diminution in the number of red corpuscles is out of proportion to the loss of hemoglobin. In making the red cell count, care should be exercised not to overlook the numerous microcytes usually present. The disproportionate loss of red cells and hemoglobin is explained by the fact that, wliile very pale corpuscles are seen, there are also many which contain a normal amount of blood pigment. Decided polychromatophiha is in- variably noted, and basophilic granulation is common. Pronounced poikilocytosis seems to develop only in the protracted cases. The presence of megalocytes and megaloblasts constitutes a particularly charac- teristic feature, and their occurrence is explained by a megaloblastic transformation of the bone marrow as far as the formation of red cells is concerned. Mi- toses of erythroblasts may also be found. These evidences of disintegration and abnormal regenera- tion of the blood are not found in chlorosis or second- ary anemia. The leucocyte count in pernicious anemia varies considerably. Leucopenia is an unfavorable prog- nostic sign. While neutrophilic leucocytosis is noted in the course of improvement, it may also be indicative of inflammatory complications. The presence of myelocytes and Tiirk's irritation forms is not uncom- mon. Plate XXII. Fig. 41 Figure 41. — Pernicious Anemia. Stained Film. Magnification 750 [Same case as Figure 37. Duration of disease four weeks.] The decided polychromatophilia and the shght poikilocytosis are apparent and probably explained by the severity and short duration of the case Lack of uniformity in the distribution of hemoglobin is also evident, some cells being very pale, while others seem to contain an excessive amount of blood pigment. Two megaloblasts with pycnotic nuclei are seen above, to the right, and in the center there is a megaloblast with a large nucleus poor in chromatin and a basophiMc cell body. A Turk's irritation form is seen immediately beneath the latter, and these two cells show a striking resemblance. A megaloblast in process of mitotic division is also seen, and to the right a polymorphonuclear neutrophile. 51 PLATE XXIII THE LEUKEMIAS The diagnosis of leukemia is based on the demon- stration of a permanent increase in the nmnber of white corpuscles in the blood, a large percentage of which are bone marrow cells or abnormal leucocytes. This statement apphes to myelogenous leukemia as well as to lymphatic leukemia, and for the present this classification seems desirable, as it is based on the condition of the blood. Further subdivisions do not seem indicated as yet, particularly as almost every case of leukemia presents individual charac- teristics. These may undergo pronounced change in the course of the disease, by variations in the relative proportion of the different varieties of cells, as well as by the disappearance of some and sudden appearance of other types. All cases show a more or less marked anemia, which may present all the characteristics of a perni- cious anemia, but some are seen, with several hundred thousand leucocytes in a cubic milhmeter, without apparent change in the red cells. The severity of the disease is appreciably dependent on the condition of the red cells. A detailed description of the changes in the blood in the different forms of leukemia will be found in connection with the following illustrations. 62 Plate XXIII. Fig. 42 Female, 23 years old. Myelogenous Leukemia. Hemoglobin, 65 per cent.; red cells, 3,200,000; leucocytes, 337,000 in 1 c.mm. Figure 42. — Myelogenous Leukemia. Fresh Double Cover-glass Specimen. Magnifi- cation 750 The decided increase in the number of leucocytes is evident. In this connection it is well to recall that some cases of leukemia may show comparatively low leucocyte counts, for example, 10,000 in 1 c.mm., whereas a leucocytosis of i50,000 or more may exist for a considerable period. Consequently the count alone is not sufficient for diagnosis. As considerable practice is necessary to differentiate the various types of leucocytes in a fresh unstained specimen, and some cannot be recognized in this waj" at all, the dried and stained films are preferable for the purpose. The red cells show no essential change. The fol- lowing four types of leucocytes can be recognized in the illustration: a. Polymorphonuclear finely granular forms, noted in Fig. 16, are neutrophiles. b. Cells of the same size with round or lobulated nuclei and coarse refractive granulation are eosino- philic myelocytes or eosinopliiles. c. Larger and lighter cells with round nuclei and finely granular protoplasm are bone marrow cells, which can be more accurately classified in the stained specimen. d. Small leucocytes of different forms which also cannot be classified in the unstained specimens. The greater the variety of leucocytes present, the more difficult the classification, in unstained speci- mens. 53 PLATE XXIV Male, 62 years old. Myelogenous Leukemia. Blood very con- centrated. Hemoglobin, 105 per cent.; red cells, 6,184,000; leucocytes, 59,300 in 1 c.mm. Figure 43. — Myelogenous Leukemia. Fresh Double Cover-glass Specimen. Magnifi- cation 750 The addition of a few drops of an aqueous solution of gentian violet to the diluting fluid differentiates the leucocytes somewhat (compare Fig. 42), but is not sufficient for the proper classification and recog- nition of the pathological forms. The red cells show but little change in shape and size. A group of blood platelets is seen in the center, on the right a poly- morphonuclear neutrophile, on the left two granular bone-marrow cells and above a lymphocyte. Figure 44. — Myelogenous Leukemia. Stained Film. Magnification 750 [The same case.] The various characteristics of the different varieties of leucocytes are apparent in the stained specimen. In the fresh specimen, for example, it is impossible to distinguish the neutrophihc myeloc>i;e from the basophilic mononuclear, as the vesicular structure of the latter looks just like the fine granulation of the former. The red cells show evidence of considerable anemia. While undue concentration of the blood has given normal hemoglobin and red cell figures as above, the anemic changes of the red cells are made apparent 64 Plate XXIV. Figs. 43, 44 in the stained specimen by differences in size, irregu- larity in shape, polychromatophiha and the presence of normoblasts. A megaloblast is seen above and a normoblast below, both showing polychromato- phiha. Two polymorphonuclear neutrophiles and blood platelets are also present. The other cells will be described later. 55 PLATE XXV CHKONIC LYMPHATIC LEUKEMIA Chronic lymphatic leukemia is characterized by the presence in the blood of varying numbers of abnormal Ij'mphocytes, usually of the small, but occasionally of the large type. The absolute counts are generally high, ranging from 50,000 to 1,000,000 in 1 c.nmi. The relative percentage of these lym- phocj^es is also high, ranging from 95 to 99 per cent, of the leucocytes, there being an absolute decrease in the other varieties. The appearance of the blood is not complex, and is absolutely characteristic of the condition. The red corpuscles are usually in fair condition, and only show extensive change in the advanced and terminal stages of the disease, when there is a de- crease in their number and a corresponding loss in the amount of hemoglobin. The blood platelets are slightly, if at all, increased in numbers. Male, 't-i years old, sick for one year. Multiple swellings of lymphatic glands. Spleen enlarged, and extending to the median line. Progressive anemia for the past three months. Duration of disease eighteen months. A complete examination of the blood could not be made, but the stained film justifies a diagnosis of chronic lymphatic leukemia. 56 Plate XXV. Fig. 45 •• • • o Figure 45. — Chronic Lymphatic Leukemia. Stained Film. Magnification 330 The increase in the number of lymphocytes is such that there are more white than red cells. The lymphocytes all appear of approximately the same size and show a very narrow margin of protoplasm, barely visible in some. There are a few somewhat larger lymphocytes with a lighter colored nucleus. An abnormal segmentation of the nuclei is noted which is foreign to lymphocytes in normal blood. (For details see Fig. 47.) The red cells show no change except diminished hemoglobin content. S7 PLATE XXVI Figure 46. — Chronic Lymphatic Leukemia. Fresh Double Cover-glass Specimen. Magnification 750 [For detailed condition of the blood see description of Fig. IS.] The nuclei of the lymphocytes are stained light blue by the addition of a drop of aqueous solution of gentian violet to the diluting fluid. Nothing ab- normal is apparent in the lymphocytes of this wet specimen, except that they appear a little larger than those in normal blood. A polymorphonuclear neu- trophile is seen on the right. The red cells show a normal amount of coloring matter. Figure 47. — Chronic Lymphatic Leukemia. Stained Film. Magnification 750 [The same specimen as in Fig. 45.] A thin smear is essential in order to recognize the abnormal features of the lymphocytes in chronic lymphatic leukemia, as they differ considerably from the small lymphocytes of normal blood. They are larger and have apparently segmented nuclei, poor in chromatin, which jjractically fill the cells and lend the impression of immaturity. There is a nar- row margin of protoplasm, sometimes barely visible. Acidophilic granules cannot be demonstrated. These cells must be considered immature lympho- cytes representing a definite stage in the development of the lymphocytes of normal blood. The red corpuscles are somewhat deficient in hemoglobin. 58 Plate XXVI. Figs. 46, 47 Plate XXVII. Fig. 48 PLATE XXVII Male, 38 years old. For past eighteen months progressive glandular swellings in the neck and both axills. At present all sui)erficial glands are palpable and about the size of a plum. Spleen 10 X 7 cm. The blood has normal appearance and con- centration. Hemoglobin, 115 per cent.; red cells, 4,928,000; leucocytes, 79,000 in 1 c.mm. Differential count: Polymor- phonuclear neutrophiles, 5.2 per cent. (4,090) ; lymphocytes, 94.2 per cent. (74,390); transitionals, 0.4 per cent. (370); eosinophiles, 0.2 per cent. (150). Diagnosis: Chronic Lymphatic Leukemia. Figure 48. — Chronic Lymphatic Leukemia. Stained Film. Magnification 330 The increase in the number of lymphocji;es is not nearly as great in this case as in Fig. 45. Most of the cells show the characteristics described above. A few of the lymphocytes have a larger amount of protoplasm, and others show an irregular or notched nucleus. On the right is seen a nuclear figure, poor in chromatin, which looks like a flattened nuclear remnant, but really has a different significance, as seen in Fig. 49. The red cells are practically normal. The plate- lets are increased in numbers. 59 PLATE XXVIII Male, 56 years old. During the past four years he has liad glandular swellings on both sides of the neck, with periods of remission. Renewed swelling has been present for the past year with the addition of glandular enlargement in both axilla, in the inguinal folds, near the eyes and at the angle of the jaw. Spleen is considerably enlarged (25 X ^^ cm.). Moderate cachexia. Hemoglobin, 105 per cent.; red cells, 5,176,000; leucocytes, 53,600 in 1 c.mm. Differential count: Polymorphonuclear neu- trophiles, 6.2 per cent, or 3,400; lymphocytes, 92.7 per cent, or 50,000, of which 16.5 per cent, or 8,900 are large fragile forms. Transitionals and eosinophiles, 0.55 per cent, each, or 300 each in 1 c.mm. Diagnosis: Chronic Lymphatic Leukemia. Figure 49. — Chronic Lymphatic Leukemia. Stained Film. Magnification 750 In addition to the abnormal lymphocytes described above, almost every case of chronic lymphatic leuke- mia shows fragmentary nuclear structures poor in chromatin. The nucleus often has the appearance of being partially dissolved, and looks hke a granular mass devoid of protojilasm. Well preserved cells of this type are rarely seen, justifying the inference that they are very fragile and non-resistant, or that they cannot exist in the peripheral circulation. Some of these resemble large lymphocytes, while others are like large homogeneous, slightly basophilic bone marrow cells. Their nuclei, poor in chromatin, are indented or lobulated, rarely showing nucleoli. These large cells must be considered as tlie large homogeneous bone marrow cells whicli enter the cir- culation at this stage of their development. In another case where the ordinary type of small 60 Plate XXVIII. Fig. 49 lymphocytes was usually present, the author noted two short periods during which the blood contained large numbers of these "fragile cells," which would seem to indicate a specific irritation of the bone marrow. When this large type of cell occurs in chronic lymphatic leukemia, it seems reasonable to conclude that there is also a lesion of the bone marrow. 61 PLATE XXIX Myelogenous Leukemia Contrary to the rather uniform changes in the blood noted in chronic lymphatic leukemia, the cases of myelogenous leukemia present a most varied and sometimes confusing picture. All types of leucocytes found in normal blood are present, in addition to myelocytes in different stages of development. The leucocyte count is always high, ranging usually be- tween 200,000 and 2,000,000, and figures below 50,000 in 1 c.mm. are rare. The leucocyte count may recede several thousand as the result of intercurrent febrile disorders (sepsis, tuberculosis, etc.), or therapeutic measures (arsenic, X-ray, etc.), but myelocytes are invariably present in the peripheral circulation, even under these cir- cumstances, and the diagnosis of myelogenous leuke- mia is evident. While there is an increase in the actual number of each variety, the polymorphonuclear neutrophile is the normal cell to which this applies chiefly The myelocytes also show high figures, and several hun- dred thousand may be present in 1 c.mm. One or other type of these bone marrow cells usually pre- dominates during the entire course of the disease, and the different types are rarely present in equal numbers. The diagnosis of myelogenous leukemia can be made on examination of the blood, even if the leuco- cyte count is not excessive. In addition to the leu- cocyte increase, these cases invariably show changes in the red cells which are, however, not as pronounced 62 as in acute leukemia. The blood platelets are al- ways decidedly increased in number. The seven plates on the following pages illustrate four types of myelogenous leukemia, each charac- teristic on account of the predominance of special cellular types. 68 Female, 25 years old. Has been ill for the past three years, and under treatment for leukemic enlargement of the spleen for two years. Occasional periods of improvement, due to adminis- tration of arsenic. On admission to hospital rather pale and emaciated. Has a greatly enlarged spleen extending to the symphysis, and numerous cutaneous thrombi. The blood is thin and coagulation is retarded. Hemoglobin, 70 per cent. ; red cells, 3,300,000 ; leucocytes, 390,000 in 1 c.mm. Differential count: Neutrophiles, 43.07 per cent., or 168,000; lymphocytes, 1.88 per cent., or 7,000; transitionals, 2.31 per cent., or 9,000; eosinophiles, 3.46 per cent., or 13,500; basophiles, 12.17 per cent, or 47,500; myelocytes, 37.11 per cent., or 145,000 in 1 c.mm. Diagnosis: Myelogenous Leukemia. Figure 50. — Myelogenous Leukemia. Stained Film. Magnification 330 The difference between the appearance of the blood in this disease and that in chronic Ipnphatic leukemia (Fig. 45) is immediately apparent. The number of leucocytes seems much greater than in the most severe leucocj-iosis. The neutrophiles, identified by their polymorphonuclear character, are in excess of other forms. Normal lymphocytes are not present in this field. Eosinophiles and basophiles are in- creased in number. The presence of leucocytes which do not occur in normal blood is the striking feature in the picture, although a low power is used. These are eosinophilic myelocytes and large cells with round nuclei and considerable protoplasm which demand a higlier power for close study. The red corpuscles are pale; a normoblast is seen above and anotlier below toward the left. Blood platelets are increased in number. 64 Plate XXIX. Fig. 50 ■ ^^^^^^^^^^^^ ■HI ■ ^ •'-•'•,"^ ^^^1 * ^ A •'^ ^^^^^^^^^^^ ^^9> n^ ' ^ <«• » ^^^^^^^^1 ••* # ^ ^' ^^^^^^^H ^ % • «£ ^ 1 ' t • ■ 1 V ^ ^ IP ^ 1 ^^' •/c 4> Hj ^ ' -* '- ^1 1 c^^H Plate XXX. Fig. 51 PLATE XXX Figure 51. — Myelogenous Leukemia. Film. Magnification 750 Stained [The same specimen.] The diversity of the cellular forms in the specimen was such that all could not be included in this com- posite picture. a. Polymorphonuclear neutrophile. h. Eosinoplnle. c. Basoph le, in eluding abnormal forms usually abundant in myelogenoua leukemia, o. Eosinophilic myelocytes. These cells vary considerably in size nave large, pale round or indented nuclei, which are frequently indistinct. lUe protoplasm is basophilic and usually distended with eosinophilic gran- ules. Some basophilic granules may be found mixed with the eosino- philic in the unusually small cells of this type. e. Homogeneous mono- nuclears with basophilic protoplasm and nuclei in different stages of development showing nucleoli. /. Homogeneous mononuclears with baso- philic protoplasm showing slight neutrophilic granulation, g. A more mature lymphocyte, h. Disintegrating eosinophile. i. Nuclear remHant with nucleolus and nuclein strands, fc. "Fragile Form." 65 A normoblast is present ( 1 ) . Polychromatophilia and basophilic granulation in red cells can be seen. There are also many blood platelets. No neutrophilic myelocytes present. 66 PLATE XXXI MYELOGENOUS LEUKEMIA The same case after the administration of arsenic and use of X-ray. Complicated by extensive tuber- culous lesion of both lungs. Two days before death. Hemoglobin, 55 per cent. ; red cells, 2,34 4,000 ; leucocytes, 78,900 in 1 c.mm. Differential count: Polymorphonuclear neu- trophiles, 56,630, or 70.42 per cent. ; lymphocytes, 9,270, or 11.75 per cent.; transitionals, 2,780, or 3.52 per cent.; basophiles, 930, or 1.20 per cent.; myelocytes, 10,390, or 13.14 per cent. Of the latter, neutrophilic myelocytes, 6,490, or 8.20 per cent. ; eosinophilic myelocytes, 190, or 0.24 per cent.; immature stages of neutrophilic myelocytes, 3,710, or 4.70 per cent. Figure 52. — Myelogenous Leukemia. Stained Film. Magnification 750 The character of the specimen has undergone a complete change. While a decided polymorpho- nuclear neutrophile leucocytosis is present, a distinct leukemic composition is still apparent. The diminu- tion in the number of eosinophiles, myelocytes and basophilic cells as compared with the former specimen is noteworthy; these having been replaced by neu- trophilic myelocytes and immature forms of this type. This change is doubtless the result of the tuberculous infection, as it was not noted during the 67 improvement caused by the administration of arsenic and the use of the X-ray. a. Polymorphonuclear neutrophilea some showing vacuoles of dege. eration (compare Fig. 15). 6. Neutrophilic myelocyte, c. Large mono- nuclears with basophilic protoplasm and numerous neutrophilic graules. d. Large homogeneous mononuclears with neutrophilic protoplasm show- ing vacuoles, e. More mature lymphocyte. The number of blood platelets has diminished. 68 Plate XXXI. Fig. 52 PLATE XXXII MYELOGENOUS LEUKEMIA Female, 23 years old. Has looked pale for past three and one- half years. Increasing enlargement of spleen for last eighteen months. At present there is decided pallor. Spleen extends 1 1 cm. to the right of median line. No glandular swellings. Hemoglobin, 65 per cent.; red cells, 3,200,000; leucocytes, 337,000 in 1 c.mm. Differential count: Polymorphonuclear neutrophiles, 94,040, or 27.9 per cent.; myelocytes, 195,930, or 58.10 per cent.; basophiles, 39,190, or 11.7 per cent.; eosino- philes, 7,840, or 2.3 per cent. Diagnosis: Myelogenous Leukemia. 69 Figure 53. — Myelogenous Leukemia. Stained Film. Magnification 750 The complex composition of the blood seen in Fig. 51 was never noted during the entire course of this case. In this specimen the leukemic change is shown by immature forms of myelocytes, instead of by typical eosinophiUc and neutrophihc myelocytes. Formerly the case would have been called a myelogen- ous leukemia with lymphoid condition of the blood. The white corpuscles are noted in groups of twenty or thirty and have been distorted by pressure. The red cells show no particular change. a. Polymorphonuclear neutrophiles. b. Eosinophiles. c. Basophilic myelocytes and basophiles. d. Mononuclears with more or loss baso- philic protoplasm. c. Homogeneous mononuclears with neutrophilic protoplasm. /. Mononuclears with basophilic protoplasm and faini. neutrophilic granulation. 70 Plate XXXII. Fig. 53 Plate XXXIII. Fig. 54 PLATE XXXIII MYELOGENOUS Li:UKEMIA The same ease after treatment with arsenic and X-ray. Hemoglobin, 90 per cent. ; red cells, 4,050,000 ; leucocytes, 14,000 in 1 c.mm. Differential count: Polymorphonuclear neutrophiles, 9,520, or 68 per cent.; myelocytes, 1,820, or 13 per cent.; basophiles, 1,260, or 9 per cent.; eosinophiles, 420 or 3 per cent. ; lymphocytes, 980, or 7 per cent. u. Polymorphonuclear neutrophiles. 6. Basophiles. c. Homogeneous mononuclears with basophilic protoplasm, d. Mononuclears with baso- philic protoplasm showing neutrophilic granulation. 71 Figure 54. — Myelogenous Leukemia. Stained Film. Magnification 750 The condition of the blood shows decided improve- ment, and considerable search is necessary before two bone marrow cells are found in the same field, as in the illustration. Owing to the persistent high percentage of myelocytes the present condition must be considered a remission rather than a cure. The bone marrow cells present show no new types. The red corpuscles show moderately diminished hemoglobin content. Some of the blood platelets are very large. 72 I PLATE XXXIV MYELOGENOUS LEUKEMIA Male, 62 years old. Suffering for the past six months from general weakness and emaciation, with enlargement of the abdo- men. Moderately anemic and cachectic. No glandular swellings. The spleen extends to the median line. Blood appears concen- trated. Hemoglobin, 105 per cent.; red cells, 5,000,000; leuco- cytes, 55,200 in 1 c.mm. Blood platelets much increased. Dif- ferential count: Polymorphonuclear neutrophiles, 38,580, or 69.88 per cent.; lymphocytes, 7,700, or 13.98 per cent.; transi- tionals, 390, or 0.71 per cent.; eosinophiles, 580, or 1.05 per cent.; basophiles, 580, or 1.05 per cent.; myelocytes, 7,350, or 13.33 per cent. Numerous normoblasts, poikilocytes, polychromatophilia and basophilic granulation of red cells. Diagnosis: Myelogenous Leukemia. Figure 55. — Myelogenous Leukemia. Stained Film. Magnification 750 During the course of the disease this case con- stantly presented a condition of the blood known as "mixed cell leukemia." There is no valid reason for retaining the term in this case, or in the one described in connection with Fig. 53. In addition to neutro- philic myelocytes, there are cells representing earlier stages of development of this type, namely, mono- nuclears with basophilic, homogeneous or granular protoplasm. This illustration and Fig. 53 show that all the forms of bone marrow cells need not necessarily be present in every case of myelogenous leukemia. Many specimens were examined, and not a single 73 eosinophilic myelocyte found. As these cells develop from the basophihc myelocytes, it was not surprising to find comparatively few basophihc myelocytes and basophiles. Subsequent developments in this case could not be followed, as the patient returned to his home. a. Polymorphonuclear neutrophile. 6. Immature lymphocyte, o. Neutrophilic myelocytes, d. Mononuclears with more or lesa marked jrianular protoplasm, e. Large mononuclears with homogeneous baso- pliilic protoplasm, f. Normoblast. Red cells show no essential change. Blood plate- lets are increased. 74 Plate XXXIV. Fig. 55 PLATE XXXV MYELOGENOUS LEUKEMIA Female, 53 years old. Confined to bed for past six months. Extreme pallor and prostration. Spleen extends to right inguinal fold. Hemoglobin, .35 per cent.; red cells, 1,843,000; leucocytes, 158,000, of which 108,000, or 68.3 per cent, are myelocytes. Many nucleated red cells, basophilic granulation, polychro- matophilia and poikilocytosis. Diagnosis: Myelogenous Leukemia. Figure 56. — Myelogenous Leukemia. Stained Film. Magnification 750 While the manifold cellular types in this picture resemble those in Fig. 51, closer study will show that they are quite dissimilar. The noteworthy feature is the presence of numerous bone marrow cells in various stages of development. The relative number of these cells is also greater than in the other cases illustrated, and probably due to the severity of the case and the absence of complicating lesions. 75 o. Polymorphonuclear neutrophiles. h. Eosinophilic myelocytes with nuclei poor in chromatin on which granules are also seen. c. Basophilic myelocytes varying in size with indented nuclei and some with densely granular protoplasm, d. Mononuclears witli basophilic protoplasm and basophilic rather than neutrophilic granulations, e. Large mononuclears with basophilic homogeneous cell body. /. Smaller cells presumably leading to the development of Ijonphocytes. g. Early stages of transi- tionals. h. Large mononuclears with neutrophilic and slightly granular protoplasm, i. Undeveloped cell with more advanced development of the protoplasm than the following, k. Parent cell. A large mononuclear witli pale homogeneous protoplasm. I. Megaloblast, showing mitosis, tlie protoplasm with basophilic granulation, to. Normoblast in the same condition. Comparing this picture with Plate I will show the relative stage of development of the different cells. The red cells show evidences of considerable anemia. 76 Plate XXXV. Fig. 56 Plate XXXVI. Fig. 57 PLATE XXXVI ACUTE LEUKEMIA The clinical picture of these cases is characterized by high temperature and a hemorrhagic diathesis, frequently combined with ulcerative conditions in the mouth and throat, and local glandular sweUing. The spleen is only moderately enlarged. The onset is acute and the general impression is that of an exceedingly severe infectious disease. A profound anemia develops rapidly and the cases invariably terminate fatally in a few days, or at most several weeks. An examination of the blood is necessary for positive diagnosis. Recent observations indicate that a condition of the blood characteristic of chronic lymphatic leukemia or of myelogenous leukemia may also be found in acute leukemia. While the changes in the blood may vary in the different cases, usually in acute leukemia an enormous number of immature bone marrow cells are found, the normal metamorphosis into more mature forms being absent. The modern view is justified that the severely acute character of the disease with leukemic changes in the blood of one or other type is pathognomonic of acute leuke- mia, the disease not necessarily demanding a specific blood picture. The cause of this rapidly fatal disease is unknown, but it would seem that the extensive ulcerations in the naso-pharynx usually present may have etiological significance. Boy, 1 1 years old. Acute onset with chills and uncontrollable epistaxis. Course of the disease characterized by high tempera- ture, hemorrhagic diathesis, profound anemia, glandular swell- 77 ings in the neck and secondary sepsis due to ulcerations in the naso-pharynx. Death at the end of three weeks. Blood decidedly hydremic. Hemoglobin, 25 to 22 per cent. ; red cells, 1,068,000 to 736,000 in 1 c.mm. ; leucocytes, 28,300 to 11,600 in 1 c.mm. Differential count: Polymorphonuclear neu- trophiles, 3.2 per cent., or 900; abnormal leucocytes, 96.8 per cent., or 27,400. No other varieties of leucocytes present. As the result of the secondary septic condition the total leucocyte count fell to 1 1 ,600, the polynuclear neutrophiles increased to 8.5 per cent., or 2,180, and the abnormal leucocytes decreased to 91.7 per cent., or 10,600. This effect of a septic complication in leukemia has frequently been noted. Diagnosis: Acute Leukemia. Sepsis. Figure 57. — Acute Leukemia. Stained Film. Magnification 750 The red corpuscles show all the usual evidences of a pernicious anemia, as follows: Decided pallor, microcytes, macrocytes, megaloblasts, poikilocytosis, polychromatophilia, and basophilic granulation. The decided diminution in the number of red cells is also apparent in the picture. The abnormal leucocytes apparently belong to one cell tj'pe, but show different stages of development. Neutrophilic and occasionally eosinophilic myelo- cytes are also found. The abnormal cells mentioned all show more or less lobulated nuclei poor in chro- matin. They have a shghtly basophilic and rarely neutrophihc protopla.sm, and many show a variously dense fine neutrophihc granulation. Vacuoles are sometimes observed. These cells are identified as immature bone marrow cells, but as they are granular, they have attained a definite stage of development ( see Plate I ) . The nuclei are decidedly indented or lobulated, and amitotic division figures in cells with a larger amount of protoplasm are frequently observed. A certain morphological change is always observed in these leucocytes, the significance of which is still obscure. A large number of the cells show proto- 78 plasmic processes of diit'erent kinds. In some a wreath of short threads surrounds the cell, while others show one or more club-shaped projections. It would seem that these are particles of protoplasm amputated from tlie cell by constrictions, and par- tially destroyed by fixation of the specimen (see Fig. 58). a. Polymorplionuck'iir neutropliile. h. Lymphocyte, c. Immature bone marrow cells, d. Neutrophilic myelocyte, e. Megaloblast. 79 PLATE XXXVII Figure 58. — Acute Leukemia. Fresh Double Cover-glass Specimen. Magnification 750 [The same case.] Gentian violet has been added to the diluting fluid. Immediately after the preparation of the specimen, a considerable number of all the leucocytes show certain globular jjrocesses as seen in the illustration. Several cyst-like structures appear attached to the cell, some being twice or three times as large as the actual cell body, the protoplasm of which is much diminished in proportion to the nucleus. The granulations are never found in the processes, but remain in what is left of the cell body- 80 Plate XXXVU. Fig. 58 Plate XXXVIII. Fig. 59 PLATE XXXVIII BLOOD CHANGES ASSOCIATED WITH TUMORS OF THE BONE MARROW At different stages of development certain tumors give rise to metastases in the bone marrow, with resulting changes in the morphological composition of the blood. In circumscribed metastases, there is only an irritation of the adjacent bone marrow, the degree of which depends on the extent of the involvement. The peripheral blood showS[ evidence of increased or hurried regeneration, due to the increased com- pensating function of the remaining healthy marrow. In other cases, there is a diffuse distribution of the metastatic process in the bone marrow, and in con- sequence the latter is largely replaced by tumor tissue. Under these circumstances the peripheral blood not only contains bone-marrow cells, but also shows a number of cellular varieties which justify conclusions in regard to the character and extent of the lesion. The red cells also undergo a more or less serious change and frequently show evidences resembling those of grave pernicious anemia. Carcinoma of the Bone Marrow Male, 33 years old. Pain in the left side after over-exertion in lifting a machine. One month later neuralgic pains also noted on the right side and in lumbar region. Progressive pallor. Clinical examination: A chain of small hard glands palpable on the right side of the neck. Rapid development of bilateral choked disk. Pleural exudate on left side. Large epitlieloid cells found in pleural and lumbar fluid. Sternum painful on pressure. Progressive anemia. Died at the end of ten weeks. Autopsy: Flat cancer of stomach a little smaller than tlie size 81 of a silver dollar, apparently quiescent. Numerous metastases in the bones (skull, ribs, vertebrae and pelvis). Blood examination day before death: Hemoglobin, lO per cent.; red cells, 1,984,000 ; leucocytes, 16,400. Differential count: Polymorphonuclear neutrophiles, 13,440, or 82.0 per cent.; lymphocytes, 1,340, or 8.2 per cent.; transitionals, 470, or 2.9 per cent.; eosinophiles, 201, or 1.2 per cent.; myelocytes, 135, or 0.7 per cent.; "abnormal cells" (tumor cells.'') 806, or 4.9 per cent. In counting 500 leucocytes, 1 20 nucleated red cells are seen, of which a moderate number are megaloblasts. Figure 59. — Carcinoma of Bone Marrow. Film. Magnification 330 St ainec In addition to a neutrophilic leucocytosis marked changes are present in the red cells, i. e., deficient A. Normoblaata. 6. Mepaloblast. c. Normoblasts with polychroma- tophilia. rf. Normoblasts showinj^ amitotic division, e. Polymorphonu- clear neutrophiles. /. Lymphocytes, g. Myelocyte, h. Abnormal cells (Tumor cells?). 82 hemoglobin, polychroinatophilia, poikilocytosis, and nucleated cells. While some of these changes are referable to the secondary anemia, the presence of an unusual num- ber of nucleated red cells is doubtless due to severe irritation of the bone marrow by the metastatic growth. The presence of munerous myelocytes in the peripheral circulation can be ascribed to the same cause, as these cells never occur in such numbers, and are frequently entirely absent in cases of secondary or primary anemia. It is, therefore, reasonable to mfer that the existing abnormal composition of the blood is referable to the active compensating func- tion of the remaining normal bone marrow, and due to direct irritation of this medullary tissue by the metastatic lesion. 88 PLATE XXXIX SARCOMA OF THE BONE MAEROW Female, 57 years old. Sarcoma of the naso-pharynx, with in- volvement of the local glands, and extension of the tumor to the base of the skull and the cranial cavity. In the early stages the case was supposedly a pseudo-leukemia. Secondary diffuse sar- comatosis of bone marrow. Death. Blood examination: Fully a year before the patient's death, certain changes in the blood, such as poikilocytosis, polychroma- tophilia, basophilic granulation, and the presence of normoblasis justified a suspicion of the participation of the bone marrow in the disease, and led to a characteristic blood picture which per- sisted for several months until the patient's death. Early in the disease the condition was as follows : Hemoglobin, 60 per cent. ; red cells, 3,104,000; leucocytes, 3,600 in 1 c.mm. Normal dif- ferential count. This was followed by progressive anemia, the appearance of megalocytes and many normoblasts, and cells of the IjTnphocyte type having larger or smaller round or indented nuclei, with more or less chromatin, and with but a narrow rim of protoplasm if any. These cells differed from the normal lympho- cyte, and from the cell seen in chronic lymphatic leukemia, and closely resembled the type of which the tumor was composed. The inference seems reasonable that these were sarcoma cells ■which had entered the circulation. At the height of the disease, eight weeks before the patient's death, the blood was as follows : Hemoglobin, 40 per cent. ; red cells, 2,608,000; leucocytes, 7,000 in 1 c.mm. Differential count: Polymorphonuclear neutrophiles, 2,300, or 32.85 per cent.; lym- phocytes, 800, or 11.42 per cent.; transitionals, 60, or 0.86 per cent. ; myelocytes, 40, or 0.57 per cent. ; tumor cells, 3,800, or 54.3 per cent. 84 Plate XXXIX. Fig. 60 Figure 60. — Sarcoma of Bone Marrow. Film. Magnification 750 Stained Composite picture. The majority of tumor cells had the appearance of (c) in the illustration, with large and frequently lobular nuclei, containing more or less chromatin and surrounded by a narrow rim of protoplasm. Abnormally large cells having large ^'esiculated or multilobular nuclei were also present as shown {d). a. Myelocytes. Me^loblaat. !>. Lfcrge ly«pk««yte«. • »*i d. Tuiii»r cells, e. 86 PLATE XL LEUKOSARCOMATOSIS Sternberg recently applied this name to a clinical condition, which will demand further observation for accurate definition. The cases may closely resemble those of acute leukemia, in the sudden development of severe constitutional disturbances with high temperatures and a hemorrhagic diathesis, resulting fatally in a few days. Concerning the gross pathology, a tumor is found in some portion of the body which has the macro- scopic appearance of a lymphosarcoma {Kundrat), or there are changes in portions of the lymphatic system having the appearance of pseudoplasm. The condition of the blood also closely resembles that usually found in acute leukemia, showing im- mense numbers of large mononuclear or multilobular abnormal cells, the average count of which exceeds 500,000 in 1 c.mm. These cells represent an atypical pathological type, which should probably not be con- sidered as a preliminary stage of the myelocyte, but rather as a "tumor cell," it being identical with the cell found in the neoplasm present, or in the tumor- hke changes of the lymphatic system. Boy, 13 years old. Has looked poorly and anemic for a con- siderable time. The illnes.s began witli pain and swelling in the throat, fever and marked prostration. On admission to the hos- pital the patient's condition was critical. He presented numer- ous ulcerations of the gums, and nuilti])le cutaneous hemorrhages about the size of a quarter, all over the body and particularly on the face, some of which were covered with scabs. The spleen wa.s undoubtedly enlarged. Death six hours after admission. Blood examination: Moderate anemia. Leucocytes, 560,000 in 1 c.mm. 86 Plate XL. Fig. 61 Autopsy : The entire mucosa of the middle and lower ileum was covered with tumors the size of a hazelnut or a walnut, resembling sarcomata, and projecting into the lumen of the bowel. Hyper- trophy of the lymphatic apparatus of the bowel, and of that at the base of the tongue. The bone marrow was grayish-red in color. Figure 61. — Leukosarcomatosis. Stained Film. Magnification 750 A certain uniformity in the cellular types charac- terizes the specimen. Unsuitably or imperfectly stained fihiis show nothing beyond many cells of similar appearance and lymphoid type. In good specimens, however, certain variations in size, con- figuration of nuclei and character of protoplasm can be made out in the cells. They all have round, more or less lobulated nuclei, often vesiculated in appear- ance, and very poor in chromatin. Several nucleoli are not vmcommon. Occasionally the nucleus is indistinct and so poor in chromatin that its outhne cannot be made out. The nuclei of the smaller cells show a denser structure and a greater amount of chromatin. The protoplasm is always scanty and often shows as a narrow margin, very few cells having shght neutrophihc granulation. The conspicuous fragmentation of the nuclei is possibly a sign of pathologically rapid growth of these cells. A certain resemblance between these cells with granulation, and the leucocytes seen in acute leukemia cannot be denied. As these cells are, however, identical with those which constitute the primary tumor, they must be considered tumor cells and not bone marrow cells. The red cells in the stained film show no essential anemic changes. An eosinophilic myelocyte is seen below to the right, and several others were found, but with this exception no other additional or pathological blood cells could be found in the entire specimen. 07 PLATE XLI BLOOD PARASITES MALARIAI. PARASITES AND TRYPANOSOMES Tertian Malaria Figure 62. — Endogenous Development of Tertian Parasite. Magnification 750 Very young parasites, just after entering the red cell, present a hyaline form. In the stained specimen a distinct ring form is seen, the brilliant red chro- matin appearing at one pole, with the so-called polar swelling directly opposite. These small tertian rings are not all equally distinct. Further development produces the "large tertian rings," particularly by growth of the polar swelling. Continued growth and irregular changes in form result in grotesque figures. The parasites are now approximately 24 hours old. An increase in the amount of chromatin, and the deposit of yellowish-black pigment are in- variably observed at this period, and the red cor- puscle, having increased to nearly or quite double its original size, is pale in color. Later the parasites appear as flattened structures, with chromatin nuclei often regularly arranged and surrounded by an achromatic zone. The chromatin nuclei increase in number, the pigment collects in a central mass or strand, and the development of the parasite is com- plete shortly before the febrile attack. It now pre- sents the chromatin granules, each surrounded by a blue ring, and the whole appearing as a regular group which constitutes the sporulation form. This now bursts and the young sporozoids are free to enter 88 other red cells to begin another 48-hour period of development. Figure 63. — Endogenous Development of Quartan Parasite. Magnification 750 The life history is very similar to that of the tertian parasite, but 72 hours are required for the develop- ment of this form. During the first day the quartan cannot be distinguished from the tertian parasite, but later it is characterized by an absence of increase in size of the affected red cell, probably due to the smaller size of the parasite. Subsequent develop- ment usually shows a band-like structm-e with a large amount of pigment instead of the grotesque forms of the tertian with less pigment. The mature parasite has a smaller number of chromatin nuclei, and the sporulation form shows ten or twelve seg- ments only, often in "sun-flower" arrangement. The small size of the quartan parasite, as compared with the tertian, . is particularly apparent in the sporulation stage. In tertian as well as quartan malaria, adult para- sites are observed in the blood, which do not resemble the ordinary forms. Some have much chromatin and a wealdy stained protoplasm, these being the male gametes. Others have a small amount of chromatin and a deeply stained protoplasm and are the female gametes. The sexual development of the parasites begins with these forms. Figure 64. — Endogenous Development of Estivo- Autumnal Parasite. Magnification 750 The endogenous development of this parasite is not perfectly understood as yet. In the early ring stage the parasites are smaller and more dehcate than the tertian or quartan, but later cannot be distinguished from the latter of the same size. Cres- cents represent the last stage in the development of 89 the estivo-autumnal parasite and change into oval or round bodies. The crescents are characteristic of estivo-autumnal malaria and are found during the febrile attacks as well as in the intervening quiescent periods, whereas the ring form is found in the febrile stage only. The crescents also develop in the red cells and are often seen surrounded by a halo of hemoglobin representing the remnant of the erythroc}i:e. In other crescents the shell of the red corpuscle is drawn hke a thread between the poles of the parasite, giving rise to basket-shaped figures. Both poles of all crescents are more deeply stained than the center, which contains chromatin and pig- ment. The gametes, or forms for the sexual develop- ment, are derived from the crescents and have the same characteristics as those of tertian or quartan malaria. 90 Plate XLI. Fig.. 62, 63, 64 "^^ r 62 '> i^ i # 9 63 • © (S) © ^ ^ %; *. ^ Plate XLII. Figs. 65, 66 PLATE XLII PARASITES OF TERTIAN MALARIA "• III Male, 28 years old. Had malaria in Algiers while servin the army. Develops a relapse after his return home. Many tertian parasites are found in tlie blood. Diagnosis: Tertian Malaria. Figure 65. — Tertian Malaria. Fresh Double Cover-glass Specimen The recognition of the malarial parasite is more difficult in fresh blood than in the stained film. Rings and young parasites with but httle pigment may escape attention, though the mature ones should be easily seen on account of the active flowing and circulating motion of their brownish-black pigment. A visible enlargement of the red cells containing the parasites in any stage cannot be made out. The parasite usually occupies the center of the red cell and abstracts its hemoglobin, which is converted into melanin, thus accounting for the pallor of the cor- puscle. In the sporulation forms, which are rarely seen, the pigment lies in the center of the occasionally rosette-shaped parasite. Two adult parasites and one sporulation form are seen in the illustration, and are easily differentiated from the leucocyte in the center. Male, 20 years old. Has always been well and never away from home. Develops chills some time after having been severely bitten by mosquitoes. The town in which patient lived contains many Italians who arrive from home each spring. The blood film shows a moderate number of tertian parasites. Diagnosis: Tertian Malaria. 91 Figure 66. — Tertian Malaria. Stained Film. Magnification 750 The specimen, obtained during the chill, shows a young tertian ring and a mature sporulation form. The ring, which shows two lateral indurations in- stead of a polar swelling, is about to enter the red corpuscle. The sporulation form still lies within the pale, scarcely visible erythrocyte. The somewhat regular arrangement of the youthful forms is evident, each chromatin nucleus being surrounded by a hght achromatic zone beheved to be nuclear juice. The pigment is seen as a slender strand. In view of the recent and moderately severe infec- tion, the red cells show no anemic changes. i 92 PLATE XLIII Schoolboy, 15 years old. Develops chills on his return from Italy. Duration of illness, six days. Numerous tertian para- sites in blood specimen. Diagnosis: Tertian Malaria. Figure 67. — Tertian Malaria. Stained Specimen. Magnification 750 The specimen was obtained a few hours before the febrile attack, and shows tertian parasites in various stages of development. After a number of attacks have occurred, the cases show so-called "sterile forms;" parasites which do not develop beyond a certain stage. If two distinct generations of parasites, maturing at different periods, are present in the blood, they give rise to the quotidian type of the disease, and present a correspondingly complex picture. The ring forms and the mature parasites shown in the illustration contain considerable pigment and but httle chromatin. The dehcate red and black granulation in the red-cell host is noteworthy, and since it has been observed with the tertian parasite only, may be characteristic of this form. Sailor, 26 years old. Has had malarial attacks for a period of eight weeks. No quinine administered. Decided anemia. Large spleen. Fever of the quotidian type. Numerous tertian parasites in the blood specimen. Diagnosis: Tertian Malaria (Quotidian). 9.3 Figure 68. — Tertian Malaria. Stained Film. Magnification 750 The specimen contains a large number of para- sites, and red cells are seen into which several ring forms have penetrated. This is beheved to be in- dicative of the estivo-autumnal type, but the type of the fever, the presence of many mature parasites, and the shght granulation of the affected erythrocytes would indicate the tertian type. The red cells show shght anemic changes. 94 I Plate XLIII. Figs. 67, 68 Plate XLIV. Figs. 69, 70 PLATE XLIV Figure 69.— Quartan Malarigi. Stained Film. Magnification 750 Three malarial parasites are seen in the field. The upper one consists of a narrow band passing through the red cell. There is a fine line of chromatin granules lying beside the body of the parasite, which is stained blue. A more mature quartan is seen on the left, in the form of a broad band with consider- able pigment and beginning increase of chromatin. The band form characterizes the quartan parasite, and in the more mature form shown in the lower part of the field, the small size, the shght degree of distortion, and the small number of chromatin granules are the differential points. Figure 70.— Estivo-Autumnal Malaria. Stained Film. Magnification 750 Two delicate rings and a crescent are seen in the field. In recent infections, and after repeated re- lapses, both rings and crescents are invariably found in the peripheral blood. In chronic malaria crescents are frequently present in considerable numbers, but the ring form is not found. In cases of malarial cachexia it may be impossible to find parasites. The small rings are more dehcate in structure than the broader ones of tertian or quartan cases. The crescents show distinct polar stain, considerable pig- ment, and are surrounded by the basket-shaped out- line of the red ceU, exceeding in size the diameter of the erythrocyte. 95 PLATE XLV TEYPANOSOMIASIS Human trypanosomiasis is due to parasites of the blood called trypanosomes, which belong to the flagellated protozoa. The parasites are actively motile in fresh blood and show the following char- acteristics in the stained film: They are fish-shaped and very slender, two or three times as long as the diameter of a red cell, with a long flagellum at the anterior extremity, and an undulating membrane on one side. A rather large nucleus with chromatin tint is found in the center. A bright red granule, the centrosome, is seen near the posterior blunt end, from which a fine line of red chromatin extends along the margin of the undulating membrane, into the flagellum. The body of the trypanosome takes a blue plasma stain. These parasites live in the plasma and not in the corpuscles of the blood, and multiply by longitudinal fission. Trypanosomiasis may take an acute or chronic course, the symptoms consisting of an irregular fever, anemia, emaciation, enlargement of the spleen and glandular swelhngs. The parasites may exist in the blood of man for years without giving rise to clinical evidences. In most cases they pass from the blood into the cerebrospinal fluid and occasion "sleeping sickness," which is a symptom of trypan- osomiasis. The disease is communicated to man by a fly (glossina palpalis), the sting introducing the parasite into the circulation. It is possible that they develop in the glossina, or they may imdergo a process of 96 Plate XLV. Fig. 71 development similar to that of the malarial parasite in the anopheles. Figure 71. — Human Tiypanosomiasis. Stained Film. Magnification Approx. 1,000 The specimen, which was obtained from a Euro- pean, contains but few trypanosomes. The lower parasite in the illustration shows a twisted body, the undulating membrane and chromatin thread, which becomes continuous with the flagellum, are plainly visible. The protoplasm of both parasites shows a shghtly granular appearance. 97 INDEX Achromatic zone, 88 Anemia, 52 in children, 43 pernicious, 47, 50 polychromatophilic, 37 primary, 47 secondary, 41 secondary, acute, 41 secondary, chronic, 42, 45 Anopheles, 97 Basophiles, 22, 24 Blood cells, human, 12 cells, human, transitional forms, 12 Blood, changes in tumors of bone marrow, 81 corpuscles, counting of, 3 development of red, 12 development of white, 12 examination of (technic), 1 examination of fresh, 8 normal, 15, 18 normal composition of, 15 normal leucocytes of, 19 normal rouleaux formation, 16 obtaining of, 1 parasites, 88 platelets, 34, 35 stain, formula of, 11 Bone marrow, blood changes in tumors of. 81 carcinoma of, 81 sarcoma of, 84 Carcinoma of bone marrow, 81 Centrosome, 96 Chlorosis, 48 Chromatin granules. 88 Coloriraetric determination, 2 Corpuscles, red, 34 red nucleated, 34 white, 19 Counting chamber, 4, 9 Diathesis, hemorrhagic, 77 Diluting fluid, 3 Diphtheria, blood changes in. 32 Eosinophile, 13, 22, 24 Erythroblast, 14, 34 Karyolytic, 35 Erythrocyte, 34 crenated, 45 polychromatophilic, 37 Estivo-autumnal parasite, endoge- nous development of, 89 Films, preparing dried, 1 staining dried, 9 Fleischl-Miescher hemometer, 2 Glossina palpalis, 96 Goicer's hemoglobinometer, 2 Granulation, basophilic, 39 Oraivitz, 14 Bagedorn needle, 1 Hayem's solution. 3 Hemocytometer, 4 Hemoglobin, estimation of the amount of, 2 Hemoglobinometer, 2 Hemometer, 2 Irritation form of Tiirck, 13 Kundrat, 86 Lead poisoning, 39 Leishman modification. 9 Leucocytes, counting the, 6 differential count of, 7, 19 Leucocytosis, 26, 32, 50 eosinophilic, 28 in child, 30 neutrophilic, 26, 29 Leucopenia. 50 Leucosarcomatosis, 86 Leukemia, 52 acute, 77 lymphatic, 52 lymphatic, chronic, 56 myelogenous, 52, 54, 62 Lymphocytes, 12, 20, 25 degeneration forms of, 25 Lymphosarcoma, 86 98 Malaria, crescents in, 95 estivo-autiimnal, 89, 95 quartan, 89, 95 rings in, 95 sterile forms, 93 tertian, 88, 91, 93 Malarial parasites, 88 Megaloblast, 13, 35 Microblast, 35 Mitotic division, 35 Mixing pipette, 4 Myelocytes, 12 Neutrophiles, polymorphonuclear, 19 Newton's rings, 5 Normoblast, 13, 34 Nuclear division figures, 35 Panoptic stain, 9 Parasites, malarial, 88 of blood, 88 Pipette, mixing, 4 PoikilocytosiB, 50 Polychromatophilia, 50 Protoplasm, basophilic, 12 neutrophilic, 12 Pseudoplasm, 86 Quartan parasites, endogenoxis development of, 89 R«d corpuscles, counting the, 4 Romanowsky method, 9 Sahli hemometer, 2 Sarcoma of bone marrow, 84 Schmalz method, 10 Sleeping sickness, 96 Specific gravity, estimation of, 10 Sporozoids, 88 Sporulation, 88 Sternberg, 86 Syphilis, infantile hereditary, 44 Tertian parasite, endogenous development of, 88 rings, 88 Thoma-Zeiss hemocytometer, 4 Toisson's solution, 4 Transitional forms, 20, 24 Trichinosis, 22 Trypanosome, 88, 96 Trypanosomiasis, 96 "Tumor cell," 86 Tiirck chambers, 3 irritation forms, 23 Weidenreich, 16 Wright's stain, 11 Zappert chambers, 3 Date Due D 000 164 456 5 WH17 SS^+lh 1920 Schleip, Karl. Hematological atlas with a description of the technic of blood examination WH17 SSUlh 1920 Schleip, Karl. Hematological atlas with a description of the technic of blood examination MEDICAL SCIENCES LIBRARY UNIVERSITY OF CALIFORNIA, IRVINE IRVINE, CALIFORNIA 92664