SARS-CoV-2 Antibody Assays May Be Unreliable More Than 30 Days After Symptom Onset

By Marilynn Larkin

November 19, 2020

NEW YORK (Reuters Health) - Assessment of SARS-CoV-2 antibody status varies among assays, which may significantly impact seroprevalence studies and convalescent plasma donations, researchers say.

"We were surprised to see such large differences in antibody detection both at early and later timepoints," Dr. Sarah Wheeler of the University of Pittsburgh Medical Center told Reuters Health by email. "We were not surprised to find low rates of non-specificity among these assays generally."

"SARS-CoV-2 antibody tests from reputable manufacturers have differing performance characteristics at different stages in antibody response and should be appropriately assessed for use case," she said. "We found that some assays detected 20-30% fewer convalescent cases than other assays, which may significantly affect seroprevalence surveys and convalescent plasma donation screening. Seroprevalence and convalescent plasma donation are two areas where sensitivity at >4 weeks post-infection is important and should be verified or clinicians and researchers may have inaccurate results."

As reported in the American Journal of Clinical Pathology, Dr. Wheeler and colleagues assessed six SARS-CoV-2 antibody assays from Beckman Coulter, Euroimmun (IgG, IgA), Roche, and Siemens (Centaur, Vista). They assessed 184 for specificity and 154 for sensitivity and seroconversion.

To assess specificity, they first tested specimens with prior coronavirus molecular positivity for possible antibody cross-reactivity. Of the 12 blood samples tested, one was collected the day after molecular testing and 11 were collected five to 28 days after molecular testing, allowing for assessment of cross-reactivity in the convalescent window. Cross-reactivity was seen only in two cases using the Euroimmun IgA assay.

Serologic assay sensitivity should be assessed, when possible, by examining antibody levels as a function of time from infection, according to the authors. "The samples available to us did not include many from patients who were symptomatic but confirmed those who were PCR negative; therefore, our assessment of assay sensitivity has inherent biases," they note.

All 69 samples used in the seroconversion assessment were also included in the sensitivity assessment. Additional samples from patients who had SARS-CoV-2 IgG and IgA testing performed as part of their clinical care (15), presurgical bloodwork (12), or for donation of convalescent plasma (58) were also included, for a total of 154 specimens.

Assay specificity was 99% or greater for all assays except the Euroimmun IgA (95%). However, sensitivity at more than 21 days from symptom onset was 84%, 95%, 72%, 98%, 67%, and 96% for Beckman Coulter, Centaur, Vista, Roche, Euroimmun IgA, and Euroimmun IgG, respectively.

The average day of seroconversion was similar between assays (8-10 days), with two patients not producing nucleocapsid antibodies during hospitalization.

The authors conclude, "Assessment of convalescent plasma donors at more than 30 days from symptom onset and seroprevalence studies should use assays with defined sensitivity at time points of interest because not all assays detected antibodies reliably at more than 30 days."

Dr. Kagya Amoako, Director, Biomaterials and Medical Device Innovation Laboratory and Founding Director and Coordinator of the Graduate Biomedical Engineering Program at the University of New Haven in Connecticut, commented by email. "It'll be great to see what a larger sample size study with appropriate controls would reveal about the variability in antibody assays performance reported," he told Reuters Health. "Using well defined assays that focus either on spike protein or nucleocapsid protein targeting, convalescent plasma donation can also be optimized for enhanced efficacy of plasma antibody therapy."

"The field may benefit from data on benchmarking of different antibody assays against molecular PCR antibody levels using a well controlled subject pool and determining the pros and cons of which antibody marker information (spike protein antibody or nucleocapsid antibodies, or both) to track for convalescent plasma therapy," he concluded.

SOURCE: American Journal of Clinical Pathology, online November 6, 2020.