Platelia sars cov 2 total ab

SARS-CoV-2 antibody positive and negative plasma used to contrive DBS specimens are described in Table 3. SARS-CoV-2 antibody positive plasma was collected from COVID-19 convalescent donors at Mount Sinai Hospital (Toronto, Canada). Plasma was tested for SARS-CoV-2 antibodies using the Platelia SARS-CoV-2 Total Ab (Bio-Rad, Hercules, California) or Anti-SARS-CoV-2 ELISA IgG (EUROIMMUN, Lübeck, Germany) kits. SARS-CoV-2 negative plasma was collected from healthy donors within the National Microbiology Laboratory (Winnipeg, Canada). To prepare plasma, blood was collected in EDTA Vacutainer tubes (Beckton Dickinson, Franklin Lates, NJ) and centrifuged at 1,500 RPM for 7 minutes. Plasma was tested using the Platelia SARS-CoV-2 Total Ab (Bio-Rad) or Anti-SARS-CoV-2 ELISA IgG (EUROIMMUN, Lübeck, Germany) kits to verify donors were negative for SARS-CoV-2 antibodies. All assays were performed according to the manufacturer’s instructions.

DBS samples were punched using a 6 mm hole punch into a 96 deep well plate. One to four 6 mm (1/4 inch) punches were added to each well and eluted in DPBS buffer (pH 7.4) containing 0.5% BSA and 0.05% Tween-20 overnight at 4°C with agitation (400 RPM). For the Anti-SARS-CoV-2 ELISA IgG (EUROIMMUN) a single punch was eluted in 500 ul of Sample Buffer overnight without agitation and 100 μL of eluate was transferred to the assay plate. For Platelia SARS-CoV-2 Total Ab (Bio-Rad) assay, two punches were eluted in 130 μL of Sample Diluent overnight with gentle agitation (400 RPM), 75 μL of eluate was mixed with conjugate, and 100 μL of the eluate-conjugate mixture was transferred to the assay plate. Elution volumes were adjusted for all other assays according to manufacturer’s specifications or in-house developed protocols (Table 4). Afterwards, plates were incubated at room temperature for 30 minutes with agitation (400 RPM) and each DBS eluate was transferred to an individual 2.0 mL screw cap tube. Eluates were stored at -80°C until shipment on dry ice to each testing site.