key: cord-1036277-2otg3lwl
authors: Anderson, Mark; Holzmayer, Vera; Vallari, Ana; Taylor, Russell; Moy, James; Cloherty, Gavin
title: Expanding access to SARS-CoV-2 IgG and IgM serologic testing using fingerstick whole blood, plasma, and rapid lateral flow assays
date: 2021-05-11
journal: J Clin Virol
DOI: 10.1016/j.jcv.2021.104855
sha: 706d2fcd8d1980feea0174c90d4e942722b129fd
doc_id: 1036277
cord_uid: 2otg3lwl

Serologic testing for SARS-CoV-2 antibodies can be used to confirm diagnosis, estimate seroprevalence, screen convalescent plasma donors, and assess vaccine efficacy. Dried blood spot (DBS) samples have been used for serology testing of various diseases in resource-limited settings. We examined the use of DBS samples and capillary blood (fingerstick) plasma collected in Microtainer tubes for SARS-CoV-2 testing with the automated Abbott ARCHITECT™ SARS-CoV-2 IgG and IgM assays and use of venous whole blood with a prototype PANBIO™ rapid point-of-care lateral flow SARS-CoV-2 IgG assay. The ARCHITECT™ SARS-CoV-2 IgG assay was initially optimized for use with DBS, venous and capillary plasma, and venous whole blood collected from patients with symptoms and PCR-confirmed COVID-19 and negative asymptomatic controls. Linearity and reproducibility was confirmed with 3 contrived DBS samples, along with sample stability and signal recovery after 14 days. ARCHITECT™ SARS-CoV-2 IgG and IgM assay results showed high concordance between fingerstick DBS and venous DBS samples, and between fingerstick DBS and venous whole blood samples (n=61). Fingerstick plasma collected in Microtainer tubes (n=109) showed 100% concordant results (R(2)=0.997) with matched patient venous plasma on the ARCHITECT™ SARS-CoV-2 IgG assay. High concordance of assay results (92.9% positive, 100% negative) was also observed for the PANBIO™ SARS-CoV-2 IgG assay compared to the ARCHITECT™ SARS-CoV-2 IgG assay run with matched venous plasma (n=61). Fingerstick DBS and plasma samples are easy and inexpensive to collect and, along with the use of rapid point-of-care testing platforms, will expand access to SARS-CoV-2 serology testing, particularly in resource-limited areas.

settings. We examined the use of DBS samples and capillary blood (fingerstick) plasma 23 collected in Microtainer tubes for SARS-CoV-2 testing with the automated Abbott 24 ARCHITECT™ SARS-CoV-2 IgG and IgM assays and use of venous whole blood with a 25 prototype PANBIO™ rapid point-of-care lateral flow SARS-CoV-2 IgG assay. The 26 ARCHITECT™ SARS-CoV-2 IgG assay was initially optimized for use with DBS, venous and 27 capillary plasma, and venous whole blood collected from patients with symptoms and PCR-28 confirmed COVID-19 and negative asymptomatic controls. Linearity and reproducibility was 29 confirmed with 3 contrived DBS samples, along with sample stability and signal recovery after 30 14 days. ARCHITECT™ SARS-CoV-2 IgG and IgM assay results showed high concordance 31 between fingerstick DBS and venous DBS samples, and between fingerstick DBS and venous 32 whole blood samples (n=61). Fingerstick plasma collected in Microtainer tubes (n=109) showed 33 100% concordant results (R 2 =0.997) with matched patient venous plasma on the 34 ARCHITECT™ SARS-CoV-2 IgG assay. High concordance of assay results (92.9% positive, 35

100% negative) was also observed for the PANBIO™ SARS-CoV-2 IgG assay compared to the 36 ARCHITECT™ SARS-CoV-2 IgG assay run with matched venous plasma (n=61). Fingerstick 37 DBS and plasma samples are easy and inexpensive to collect and, along with the use of rapid 38 point-of-care testing platforms, will expand access to SARS-CoV-2 serology testing, particularly 39 in resource-limited areas. 40

Serologic testing for SARS-CoV-2 antibodies can be used to complement PCR-based 44 diagnostic testing [1] and may assist in identifying asymptomatic cases or those with past 45 infection as potential donors for convalescent plasma therapy. The COVID-19 pandemic has 46 presented unique barriers to achieving widespread serologic testing, such as logistical and 47 4 need for venipuncture. Previous studies found no difference in IgG serologic assay results when 65 using capillary versus venous plasma [6, 7] . 66

In this study, we assessed the feasibility of using DBS and capillary plasma as the starting 67 material for the Abbott ARCHITECT™ SARS-CoV-2 IgG assay, approved under Emergency 68

Use Authorization (EUA) to detect IgG antibodies against the SARS-CoV-2 nucleocapsid protein 69 in human serum and plasma, and a prototype ARCHITECT SARS-CoV-2 IgM assay that 70 detects IgM antibodies against the spike protein. We also examined clinical performance of 71 these assays using DBS generated from venous or capillary whole blood compared to plasma. 72

Finally, we performed a preliminary evaluation of the Abbott PANBIO™ lateral flow SARS-CoV-73 2 IgG assay performance compared to the ARCHITECT SARS-CoV-2 IgG assay. 74

Contrived DBS samples were used to optimize ARCHITECT™ SARS-CoV-2 IgG assay 78 parameters, followed by a clinical performance study (IRB# 20041610-IRB01) cohort without COVID-19 symptoms was also collected. After providing informed consent, 89 venous blood was collected into vacutainer tubes and fingerstick blood was collected in 90 microtainer tubes and tested with the ARCHITECT SARS-CoV-2 IgG assay to obtain 91 comparative Index results. 92 PANBIO lateral flow SARS-CoV-2 IgG assay performance using whole blood was compared to 93 the ARCHITECT SARS-CoV-2 IgG assay run with matched plasma using samples collected as 94 

After providing informed consent, venous (12 ml) and fingerstick blood was collected from each 103 study participant (n=61). Five DBS samples each were generated from fingerstick and venous 104 blood. A 6 ml aliquot of venous blood was frozen and stored at -80°C and the remainder was 105 processed into plasma. Deidentified plasma, whole blood, and DBS samples were shipped to 106

Abbott Diagnostics (Abbott Park, IL) on dry ice. DBS results from IgG and IgM assays were 107 compared to matched plasma to evaluate clinical performance of the sample type. 108

All samples were run on an Abbott ARCHITECT i2000SR instrument using the EUA-approved 

Assay volume optimization and stability studies used venous plasma from commercially 121 available SARS-CoV-2 IgG-positive patients which were serially diluted in negative whole blood. 122 DBS samples were generated by pipetting 70 µL of each blood dilution to the center of a 12-mm 123

Whatman 903 (GE Healthcare/LabMate) DBS card (5 replicate spots/card) and dried overnight. 124 DBS were punched into 1.5-mL Eppendorf tubes to which 300 µL elution buffer (1X PBS pH 7.4, 125 0.25% Triton X-100) was added. Samples were placed on a room temperature shaker for 1 126 hour, and eluate was transferred into 2mL cryogenic tube. Samples were centrifuged for 2 127 minutes at 10,000 RCF before running on the ARCHITECT i2000SR using the SARS-CoV-2 128

IgG or IgM assays. 129

and venous blood DBS samples from 4 study participants were tested in triplicate to assess 135 reproducibility. 136

A commercially available SARS-CoV-2 IgG positive plasma sample (Sample 1) was serially 140 diluted into negative whole blood for DBS production. DBS were eluted and run with modified 141 SARS-CoV-2 IgG assay parameters to assess recovery at different sample volume inputs 142 at room temperature (RT), -20°C, and 37°C and at 1, 3, 7, 10, and 14-day intervals ( Table 2 ). Of note, discordant results occurred in 2 participants who each had 186 previously tested positive for SARS-CoV-2 by both PCR and SARS-CoV-2 IgG. At the time of 187 sample collection for this study, these 2 participants had begun to serorevert and their SARS- Table 3 ). Like the 213 SARS-CoV-2 IgG assay, the single discordant venous DBS sample occurred in a seroreverting 214 patient whose SARS-CoV-2 IgM plasma index was 0.96, near the assay cutoff. 215 

Qualitative SARS-CoV-2 IgG reactivity from each of the 61 DBS study participants was 224 determined using venous whole blood with the prototype PANBIO lateral flow SARS-CoV-2 225 assay (Abbott Rapid Diagnostics Jena GmbH). ARCHITECT SARS-CoV-2 IgG concordance 226 with the PANBIO interpretation was 26/28 (92.9%) and 33/33 (100%), respectively (Table 2) . 227

One of the two discordant samples was from a study participant who had an ARCHITECT index 228 of 1.46, which is near the assay cutoff. 

A total of 109 participants provided informed consent to have venous blood drawn and receive a 236 fingerstick to generate matched plasma samples collected in Vacutainer and Microtainer tubes, 237 respectively. One participant was excluded due to insufficient fingerstick plasma collected. We investigated different ways to expand and improve access to SARS-CoV-2 antibody testing 253 using sample types that require less processing and handling than current methods requiring 254 trained phlebotomists. Our results confirm the feasibility of using DBS for SARS-CoV-2 IgG and 255

IgM detection and showed good concordance with plasma index values on the ARCHITECT 256 SARS-CoV-2 IgG and IgM assays. Furthermore, the robust reproducibility and equivalence 257 observed between fingerstick and venous DBS suggests that this method could be utilized in 258 routine clinical testing as an alternative to current venous plasma. SARS-CoV-2 IgG DBS had 259 higher background in known negative samples, but the results were below the index threshold. 260

Future experiments with different elution conditions will be conducted to further optimize 261 background reduction. Importantly, DBS generated from venous and fingerstick blood produced 262 concordant assay results and were stable for 2 weeks at room temperature (<±10% change). 263

This observed stability has important implications for the potential use of DBS in large field 264 collection studies in low income geographies where access to freezer storage may be limited or 265 unavailable and these results highlight the benefit that DBS could bring to such collection 266 studies. Notably, two participants had negative IgG results with plasma but positive results with 267 DBS. These 2 participants with discordant IgG results and one participant with discordant IgM 268 results had previous positive PCR and IgG results and were found to be in the process of 269 Finally, we have shown good concordance between ARCHITECT SARS-CoV-2 IgG assay run 292 with plasma and the PANBIO rapid point-of-care lateral flow SARS-CoV-2 IgG assay using 293 blood. The ability to rapidly, accurately, and affordably determine seroprevalence in a population will be an important tool in the growing arsenal of SARS-CoV-2 diagnostic testing, particularly in 295 resource-limited areas. The simplicity of performing the lateral flow assay with whole blood also 296 eliminates the need for centrifugation and plasma separation steps, further reducing cost and 297 complexity of obtaining a result. 298

Expanding access to SARS-CoV-2 antibody testing will likely require combinations of different 300 testing methods. We have shown that expanding testing capabilities using DBS, Microtainers, 301

and rapid point-of-care tests is feasible and that results delivered with these methods are 302 comparable to current testing approaches. 

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