key: cord-0930392-bkpbzbaw
authors: England, J. J.; Riegel, C.; Jankowski, L. D.; Todd, W. J.
title: A micro cell culture method utilizing modified microscope slides for use in fluorescent antibody and enzyme immunoassays
date: 1982
journal: J Tissue Cult Methods
DOI: 10.1007/bf01666874
sha: 1874f0438ef35b4677b5eb16bbb08abfa42023ac
doc_id: 930392
cord_uid: bkpbzbaw

A technique for culturing small quantities of mammalian cells on modified microscope slides is described. The modified microscope slides were Bellco Glass, Inc., toxoplasmosis slides and the cell cultures used were early passage bovine embryonic lung cells and continuous cell lines of porcine and canine origins. The slide cell cultures were either uninfected or infected with selected viruses or the obligate intracellular protozoanEncephalitozoon caniculi for utilization in direct and indirect fluorescent antibody testing or in peroxidase antiperoxidase immunosorbant assays.

Mammalian cell cultures infected with viruses, or other microorganisms, are utilized routinely as substrates for direct or indirect fluorescent antibody studies (1, 2) or for enzyme immunoassay (3, 4) . CeU cultures for such purposes have been prepared in Leighton tubes, chamber slides, or microtiter plates; however, these systems require moderate to large quantities (0.2-2 ml) of reagents and are cumbersome to manipulate. Described herein is an inexpensive and simple micro cell culture technique that utilizes commercially available toxoplasmosis microscope slides; these slides have eight circumscribed areas (each six mm in diameter) which hold 0.025 ml of fluid. Additionally, these slides can be prepared en masse and easily stored for later use. The cell culture systems utilized in this study all grew satisfactorily on the toxoplasmosis slides. Continuous cell lines grew more efficiently than low passage cells. Users of this technique must utilize a humid chamber and either a CO2 incubator or HEPES buffer in the culture fluid to prohibit evaporation or pH change since each circumscribed area holds only 0.025 ml of fluid. The configuration of the toxoplasmosis slides permits the use of at least six wells for test purposes and one area each for positive and negative controls.

Use of these systems for EIA tests has shown considerable promise since serial dilutions may be used and since controls for each slide are also present. Of special interest was the ease with which these slides may be developed en masse using Jerne racks, thus facilitating the use of minimal amounts of reagents with uniformity of reaction time for the peroxidase techniques. Counterstaining with hematoxylin greatly facilitates reading EIA slides by providing a contrast between the nucleus and cytoplasm of the cells (Figs. 3, 4) .

Use of these slide cultures for fluorescent antibody testing permitted the preparation of stock cultures for use as needed. These slides also may be rinsed using conventional hema-tology wash racks, and they may be easily cover slipped using FA mounting medium for FA examination. In addition, the slides used herein were of sufficient quality to permit direct examination with fluorescent microscope systems.

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