Efficacy of POC Antibody Assays After COVID-19 Infection and Potential Utility for "Immunity Passports"

Akram Shalaby, MD; Hansini Laharwani, MD; John T. Bates, PhD; Patrick B. Kyle, PhD

Disclosures

Lab Med. 2022;53(3):262-265. 

In This Article

Materials and Methods

Sixty previously tested patient specimens designated for disposal were obtained from the University of Mississippi Medical Center (UMMC) laboratory for this evaluation. Each specimen was collected from patients who presented to UMMC with symptoms suspicious for COVID-19 from late March to mid-April 2020. Of the 60 specimens, 30 originated from individuals positive for SARS-CoV-2 and 30 from negative individuals as determined by RT-PCR using the Abbott RealTime SARS-CoV-2 on an Abbott M2000 analyzer. The serum specimens were collected at a mean of 13.4 days after symptom onset (range, 7–30 days; lower quartile, 9.75; upper quartile, 15.5). The mean age of all patients included in the study was 54 years (range, 5–95 years). Thirty-one of the patients in the study were males and 29 were females. See Table 1 and Table 2 for patient demographics.

BioMedomics and Premier Biotech Rapid IgG-IgM Antibody Assays

Two commercially available point-of-care lateral flow assays manufactured in China and distributed in the United States by BioMedomics (Morrisville, NC) and Premier Biotech (Minneapolis, MN) were evaluated. Each of the assays are qualitative in nature and are designed to detect IgG and IgM antibodies specific to SARS-CoV-2 in serum, plasma, or whole blood. Package inserts do not state the antigen(s) used in either test or the expression systems used to generate the antigens. Each test cassette was received sealed in a foil pouch with a desiccant and buffer. An alcohol pad and lancet were also provided with each Premier Biotech test cassette. Each test cassette has 3 regions that contain reaction antigen for IgG antibodies, IgM antibodies, and a positive control. The presence of antibodies is indicated by the appearance of a purple line in the IgM or IgG regions, which indicates a functioning test. Testing was performed as per each manufacturer's instructions. We added 10 μL of serum to the sample well in the respective device, followed by 2 drops of buffer solution in the buffer well. Results were determined visually after 15 minutes had elapsed.

IgM and IgG ELISA Assays

Per Stadlbauer et al,[9] ELISAs were developed for the measurement of human IgM and IgG antibodies specific for the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. We coated 384-well MaxiSorp plates (Thermo Fisher Scientific) with purified recombinant RBD at a concentration of 3 μg/mL in phosphate-buffered saline (PBS). Recombinant RBD that was produced in Sf9 insect cells was purchased from Genescript. The coating volume and reaction volumes were 25 μL per well. Plates were incubated overnight at 4°C, washed 3 times with PBS containing 0.1% Tween20, and blocked with PBS containing 3% dry milk for 1 hour at room temperature. The blocking buffer was removed, and specimens (1-log dilutions in blocking buffer, 5 × 101-5 × 104) were added to the wells. Plates were incubated for 2 hours at room temperature before washing 3 times. Horseradish peroxidase conjugated to anti-human IgG FC (Southern Biotech) or anti-human IgM (Southern Biotech) was diluted 1:3000 in PBS containing 1% dry milk, added to the wells, and incubated at room temperature for 1 hour. The plates were washed 5 times and developed with tetramethyl-benzidine (Southern Biotech). After 30 minutes, development was stopped by adding 25 μL of 2N H2SO4 to each well. Absorbance was measured at 450 nm. The endpoint dilution titer was set to the serum dilution that resulted in an absorbance of 0.2 absorbance units over background. A specimen was counted as having a positive result if the reciprocal of the endpoint dilution was >5000 absorbance units.

Abbott SARS-CoV-2 IgG Assay

The SARS-CoV-2 IgG assay (Abbott Diagnostics, Abbott Park, IL) is a chemiluminescent microparticle immunoassay designed for the detection of IgG antibodies to the nucleocapsid protein of SARS-CoV-2. For this assay, 150 μL of serum or plasma is mixed with SARS-CoV-2 antigen-coated microparticles and allowed to react. After a wash step, anti-human IgG acridinium-labeled conjugate is added, is allowed to incubate, and is washed. Chemiluminescence is measured as relative light units. The presence of antibodies is associated with increasing luminescence. A cutoff of 1.4 S/C was used for positivity as per the manufacturer. All analyses were performed on an Architect i2000SR (Abbott Diagnostics) after proper calibration as recommended by the assay manufacturer.

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