Expires: June 8, 2023
Elitza Theel, Ph.D.
Associate Professor of Laboratory Medicine and Pathology
Director, Infectious Disease Serology Laboratory
Mayo Clinic, Rochester, Minnesota
Contact us: email@example.com.
Hello and thank you for joining us today for this week's Hot Topic on serologic testing for IgG antibodies against SARS-CoV-2. My name is Elli Theel and I direct the Infectious Diseases Serology Laboratory at Mayo Clinic in Mayo Clinic Laboratories.
Before we start just to note that I do not have any relevant disclosures to share.
Alright, so just a little bit of background on antibody development following infection with SARS-CoV-2: as we all know, because this is a new virus to which no one has been exposed in the past, after infection it takes some time for virus-specific antibodies to develop, and that's what's being shown here in this graph. As you can see, after symptom onset, it takes a good one to two weeks for antibodies to develop, and this delay is why we want to rely on molecular tests, not antibody tests to diagnose acute infection early on.
Although we are still learning about our immune response to the virus, we know that most individuals will be antibody positive after two weeks of symptoms, and while IgM-class antibodies seem to decline about five to seven weeks later, IgG-class antibodies will remain detectable for much longer. Given this understanding of the timing of our immune response, what role do serologic tests play for SARS-CoV-2 in the current environment?
There's general consensus among multiple organizations that serologic testing is currently useful for the purposes of epidemiologic or prevalent studies and potentially to facilitate contact tracing. Additionally, antibody tests are necessary for identification of potential convalescent plasma donors and will be essential to evaluate the efficacy of candidate vaccines as they enter clinical trials. And then finally, these tests may be used as potential aides for the diagnosis of COVID-19 in patients who are negative by a molecular test and who present later on in their disease course.
There are also scenarios for when antibody testing is not recommended, including as mentioned earlier for diagnosis of acute COVID-19. They should also not be used to determine whether or not a patient has developed protective immunity against reinfection. And therefore because of that, unknown antibody results should not be used to guide the use of personal protective equipment or adherence to social distancing practices or recommendations.
Right now there are about 190 commercial serologic tests for SARS-CoV-2 that are available, and they’re shown here below. Of these, 12 have received emergency use authorization from the FDA, as of today. Also, the FDA has announced that there are 29 serologic tests whose manufacturers did not submit for or receive emergency use authorization, so they recommend that those tests no longer be distributed or used, and that list can be found on the FDA website.
The serologic tests that are available differ in their design, including in their format, which can for the most part be separated as either rapid lateral flow assays or more complicated enzyme-linked immunosorbent assays or chemiluminescent-based immunoassays. These tests also differ in what type of antibody is detected and on what SARS-CoV-2 viral protein is used in the test kit. I've listed the three most common proteins used here. So my point here is that there's quite a bit of variety between the serologic tests available, and because of these differences, as you can imagine, they do not all perform identically to each other.
This is just a list of the 12 serologic tests that currently have emergency use authorization. Again, this is to show you that these vary from the sample type that's accepted to their overall design. Currently for our Mayo Clinic Laboratories clients, we're offering the Ortho Clinical Diagnostics, Anti SARS-CoV-2 IgG chemiluminescent assay. The obvious question that comes up is, how well does this test perform to detect IgG antibodies to SARS-CoV-2?
To briefly summarize our verification studies for this test, to look at sensitivity we collected serial serum samples from COVID-19 confirmed patients and evaluated them by the Ortho test. And as you can see, patients start to seroconvert between 9 and 12 days post-symptom onset. But after about 14 days of symptoms, all individuals seroconvert to IgG positive, and you can see that shown in both of the graphs below.
We also looked at specificity using samples collected prior to 2019 and in serum samples which were positive for antibodies to other infectious pathogens as well. And as you can see in the table on the top, we found an overall specificity rate of approximately 99.6%, which is consistent with what the manufacturer reported to the FDA. In addition to sensitivity and specificity, however, we also looked at the positive predictive value of a test, or of this test, I should say, which determines what the probability is that an individual who tests positive by the test truly has antibodies to the virus. And this probability is impacted by both the specificity of the assay but also by the prevalence of the disease. And in general, the lower the prevalence of the disease, the lower the positive predictive value of that test, meaning that there's a higher risk of a positive result occurring in an individual who has not been infected. But without going into the calculation details, you can see that if we use the Ortho test—which, again, has excellent sensitivity and specificity—but if we use this in a region where prevalence is 1%, the positive predictive value is about 72% compared to using the same test in a region where COVID-19 prevalence is 5%, and we see that the positive predictive value jumps to 93%. So you can see why it's really important to use these tests wisely and to also try to use the most specific test possible.
When it comes to serologic test interpretation, generally a negative antibody result indicates no prior infection or exposure to the virus with the caveat that results will be negative if samples are collected too soon following infection or they're collected from an individual who is significantly immunosuppressed. A positive result, on the other hand, suggests that the patient was infected at some point in the recent or more distant past, as we talked about just now, though the accuracy of this result will really be impacted by the local prevalence of the infection and the overall specificity of the test itself. What we need to keep in mind also is what these results do not yet tell us, which is whether or not a patient with a positive antibody result is actually protected against reinfection. And because of this unknown, we cannot yet use a positive antibody result to guide decisions regarding adherence to social distancing recommendations, or the use of masks or other personal protective equipment.
So what do we know about protective immunity against reinfection with this virus? As we all know, our immune response and immunity itself is multifaceted and not solely dependent on antibodies. That being said, IgG class antibodies do play a big role in protective immunity, and it's specifically IgG neutralizing antibodies that are key. Briefly neutralizing antibodies inhibit the virus from infecting other cells, but unfortunately not all antibodies are neutralizing and the commercially available tests that we currently have do not distinguish neutralizing from non-neutralizing antibodies. Testing for neutralizing antibodies is challenging for various reasons that I won't get into, but that is why they are not yet widely available in the U.S. That being said, we know from initial studies that most individuals will develop neutralizing antibodies after infection and we know that reinfection with SARS-CoV-2 does not seem to occur in preliminary studies with rhesus macaque monkeys, which alongside other incidental findings does seem to suggest that we will likely generate at least some immunity to reinfection for at least a short period of time. But the question remains: what level of neutralizing antibodies is protective and how long does that protective immunity last?
In summary, I think it's a great stride forward that we have increasing oversight for the available serologic tests that are currently on the market through the FDA and the EUA process. I think the role of antibody testing will evolve over time as we gain a better understanding of our immune response to SARS-CoV-2, but currently it's most useful for epidemiologic studies, convalescent plasma donor screening, and vaccine clinical trials. I shared with you some of the performance characteristics for the serologic assay that we're currently using, which has emergency use authorization. And again, I want to underscore the importance of being very strategic and smart about how we use these assays, particularly in low-prevalence settings. We also talked about the importance of neutralizing antibodies, which are able to independently inactivate the virus leading to immunity, but they are not the only component of our immune response that's important. And then finally, I think the big unknown right now is what level and duration of protective immunity against reinfection is provided following an initial infection with SARS-CoV-2. And once we have a better sense of this answer, I do think that antibody testing will have a much bigger role to play in our effort to control this pandemic.
Thank you for your attention.