As the COVID-19 pandemic evolved over the past year, so too did available antibody tests for SARS-CoV-2 and clinicians’ understanding of how to use them. Throughout this time, AACC played an active role in helping the field to navigate challenges, forming a COVID-19 task force last spring to tackle ever-increasing questions on the utilities of serology testing, said Y. Victoria Zhang, PhD, MBA, vice chair for clinical enterprise strategy and director of clinical chemistry for the University of Rochester Medical Center, in New York. The group published initial recommendations in 2020 about the use and limitations of serology testing but felt there was a need for a more in-depth guideline.

This spring, the expert panel—including professionals from clinical chemistry, microbiology, and immunology laboratories, as well as specialists from in vitro diagnostics, industry, and regulatory agencies—released a document of practical recommendations for implementing and interpreting evolving SARS-CoV-2 emergency use authorization (EUA) and laboratory-developed test (LDT) serologic testing. The recommendations were published in Clinical Chemistry (Clin Chem 2021; doi:10.1093/clinchem/hvab051), with a longer, more in-depth document available on AACC’s website. “The performance characteristics of these assays and their clinical utility continue to be defined in real-time,” cautioned Zhang, the publication’s lead author.

The document provides information about serologic assay design, antibody classes, and the kinetics of the humoral immune response, as well as verification and validation of both EUA and LDTs. It also discusses four indications for serologic testing: supporting the diagnosis of COVID-19 and related sequelae (e.g., multisystem inflammatory syndrome in children); identifying potential convalescent plasma donors and manufacturing of convalescent plasma; epidemiologic and seroprevalence studies; and vaccine efficacy studies.

“Though various organizations have published guidelines on clinical utilities of serology testing, ours is the first and most comprehensive document we know so far for the implementation of the tests, particularly EUA tests, in clinical laboratories,” Zhang said. “We wanted this guideline to be a reference for our own labs and trainees for implementing EUA and LDT serology tests in-house. It is our intention to provide a full reference for laboratory professionals and healthcare workers to appropriately implement these assays in the clinical laboratory and interpret the results to serve their patient needs during this pandemic.”

Evaluating SARS-CoV-2 Serology Assays

The team went to work with the goal of providing better guidance for clinical laboratories on what to consider when bringing in a serologic test and how to validate and implement it, said coauthor Elitza Theel, PhD, director of the infectious diseases serology laboratory at Mayo Clinic and a professor of laboratory medicine and pathology. This included “everything from how serologic testing should be used, to how many samples to consider for accuracy studies, and what to think about for specificity studies, etc.”

They also designed the document to serve as a useful resource for future disease outbreaks. “This probably won’t be the last pandemic. There was Zika virus and dengue, so we designed this as a broader guidance for things to consider when implementing serologic assays for emerging pathogens in the future,” Theel said.

Clinical laboratories can find multiple uses for the document, Zhang noted. It could be a reference guide for understanding the host immune response to SARS-CoV-2, antibody kinetics, and available EUA assays; or it could help laboratorians understand the clinical utilities and limitations of serology testing and gain insights into the nuances in implementing EUA or LDT serology testing.

Importantly, the authors say, serology tests are not recommended for diagnostic purposes for SARS-CoV-2 infection. But they could identify different types of antibodies, such as IgG and IgM, and antibodies for S and N proteins or neutralizing antibodies. There are a lot of vaccine-preventable diseases for which laboratories perform serologic testing, Theel said, but the key is first identifying a minimum antibody threshold or correlate of protective immunity for those pathogens.

“I can see serologic testing playing a much more prominent role for COVID-19, maybe to guide potential future boosters or revaccination policies,” she said. “But until that happens, the role of serologic testing is going to be pretty minimal.”

How to Handle Questions on Utility and Interpretation

The guidance is timely. Clinicians continue to look to laboratories for answers, Theel said, most commonly now questioning why their vaccinated patient tested antibody-negative, and what that means. “We start by saying, ‘You don’t need to test,’ and ‘Did you order the right test?’ because with an assay ordered after vaccination, a patient’s result is going to be negative unless they were previously infected,” she said. “We also talk about whether [the patient is] immunosuppressed, because we are seeing that immunosuppressed individuals are not as reactive.”

Clinicians also are using antibody testing to see if people have been infected without knowing or to confirm the course of disease for people that are known to be positive or to have been positive, said Robert Boorstein, MD, PhD, medical director for Lenco Diagnostic Laboratories in New York City, a large reference laboratory.

“One has reason to be somewhat optimistic that the number of new infections is going to continue to decrease in the U.S., so the need for monitoring current infection presumably is going to go down as well,” Boorstein said. “As more people become vaccinated, the utility will largely be in helping determine how long vaccination lasts. The question is whether that information will be useful in aggregate, or for individual patients.”

A major push in this area is determining the immune correlate, said Debra Poutsiaka, MD, PhD, an infectious disease physician and vice chief for clinical affairs for the Division of Geographic Medicine and Infectious Diseases at Tufts Medical Center, in Boston, who agrees with Theel. Serologic tests for now don’t indicate whether a person is or is not protected from getting symptomatic or severe COVID-19, she said, making their clinical utility in some cases “essentially nil.”

“There’s not a lot that we can actually do with confidence based on a test result, whether it’s negative or positive,” Poutsiaka said, noting that her infectious disease colleagues rarely send samples for antibody testing for COVID-19. “That’s mainly because there’s no clinical action that should be dictated by the results of that test, because there is no data to back up a clinical action.”

Multiple arms of the immune system can combat COVID-19, Poutsiaka added, and antibody testing is not going to identify some of these other means of protection. For example, there is an arm of the immune system called cellular immunity, which is separate from antibody-mediated immunity.

“Multiple studies have demonstrated that vaccines and natural infection generate cellular immunity against COVID-19, but this will not be measured by antibody tests,” she said. “Right now, we’re not close to having a global assessment of immune protection from COVID-19, so I think studies will be moving in that direction.”

Research Needed to Standardize Assays, Link Results to Protection

As vaccination rates climb in the U.S., there is some interest in studying antibodies to the N protein, Boorstein said, and being able to distinguish between people who have been infected and those who have been vaccinated. Theel and Poutsiaka said there also is a need for standardization among assays—across institutions or even across countries—because it is difficult to know if antibodies found in one laboratory will translate to a patient at another location. A test, methodology, and patient population can differ among locations, Poutsiaka noted.

“Clinical tests we send out are essentially qualitative, and some are semiquantitative, but basically it’s a yes or no,” she said. “It’s hard for us to know how yes is a yes, how positive is that yes, how high is the antibody level, and even if we knew that, we don’t know yet if that translates into protection. That’s the critical link—to get an idea of whether or not someone is protected against COVID-19 based on their antibody tests. That absolutely doesn’t exist right now.”

One recent study (Nat Med 2021; doi:10/1038/s41591-021-01377-8) pulled data on observed protection from seven current vaccines and convalescent cohorts and did some statistical modeling to try to determine how antibody levels would translate to protection against symptomatic and severe SARS-CoV-2 infection, Poutsiaka said. That’s the type of work that will be “very valuable for providing a framework for how to standardize or think about measuring antibody levels, and how to figure out what’s protective and what’s not,” she said.

Meanwhile, Zhang said, tests continue to evolve from qualitative to semiquantitative to quantitative: “The more information we can get from one test, the better we will understand the science, and understand the clinical utilities of serology testing in serving patient needs.”

Karen Blum is a freelance medical and science writer who lives in Owings Mills, Maryland. +Email: [email protected].