![]() ![]() Positive patients were previously diagnosed with COVID-19 by RT-PCR or TMA (Transcription-mediated amplification) methods at our institution, or at other in the Texas Medical Center (Baylor St. Specimens for our validation study were obtained from healthy volunteers and from confirmed COVID-19 patients, under a protocol approved by the Baylor College of Medicine Institutional Review Board. ![]() Mean absorbance readings for each of the calibrators is plotted along the y-axis versus the calibrator concentrations in AU/ml along the x-axis and calibration curve is achieved using a linear regression curve-fit. The manufacturer provides a set of 3 calibrators and three levels of controls. Samples with concentration 12 AU/ml are considered reactive and samples with AU/ml 10–12 are considered equivocal. The IgG and IgM ELISA assays are semiquantitative, and report measurements in standard arbitrary units (AU/ml). The SARS-CoV2 IgG assay uses SARS-CoV-2 recombinant proteins, targeting antibodies which recognize epitopes of the nucleocapsid (N) and spike (S) proteins, whereas the SARS-CoV2 IgM ELISA uses anti-human IgM capture antibody. The SARS-CoV2 IgG and SARS-CoV2 IgM ELISA Immunoassays (Ansh Laboratories) were evaluated for use on the Dynex-DS2 automated immunoassay system. The ELISA assays target different epitopes of SARS-CoV-2 and permit distinction of antibody subtypes. ![]() To minimize false positive test results from the use of a single assay, and to further adhere to CDC’s recommendation of orthogonal testing algorithm, we validated IgG and IgM ELISA immunoassays for use as confirmatory testing. Recently our laboratory successfully validated and implemented a total anti-SARS-CoV-2 antibody test (CoV2T) on the Vitros 5600 automated chemistry analyzer. The increase in test specificity offered by the combination of two tests rises significantly when the viral antigen targeted of the two tests are distinct. Additionally, to reduce the likelihood of a false-positive result and maximize the positive predictive value of testing, the CDC Interim Guidelines for COVID-19 Antibody Testing suggests an orthogonal testing algorithm so that individuals who are positive by one antibody test are retested with a second antibody test. To assure the quality of the available tests, as of May 4, 2020, the FDA has required commercially marketed serologic tests for SARS-CoV-2 to receive Emergency Use Authorization (EUA). Studies suggest that the average time to seroconversion for IgM and IgG antibodies is 13 days after onset of symptoms, however, the titer or type of antibodies that confer protection are not yet established. The absence of recurrent cases of COVID-19 so far, and the success of convalescent plasma treatment in many cases, suggests that patients infected with SARS-CoV-2 may produce neutralizing antibodies against the virus. However, it remains unknown for how long IgG or IgM antibodies to SARS-CoV-2 remain present in circulation after the infection has been cleared. IgM antibodies are known to develop earlier in infected patients and are most useful for determining acute infection, whereas IgG may not develop until later but remain present for a longer period of time. Currently available serological tests for SARS-CoV-2 measure IgG, IgM, IgA or a combination of this antibodies. In response to the urgent need for reliable antibody detection, there has been a rapid development in serological assays for SARS-Cov-2. Serological tests are routinely used for diagnosis and management of many viral diseases to verify that an individual has had exposure to a pathogen and mounted an immune response. Recent studies suggest that combining RNA and antibody testing improves the sensitivity of diagnosis in COVID-19 patients in different phases of the disease, and provides a way to determine a past infection. Moreover, in individuals who have recovered from COVID-19, a negative RT-PCR result provides no information about prior exposure. However, viral loads in upper respiratory tract secretions peak early during disease course and may quickly decline below the limit of detection for patients presenting later in the course of infection. Laboratory diagnosis of SARS-CoV-2 infection is primarily based on viral RNA detection via RT-PCR. Timely and accurate diagnosis of the SARS-CoV-2 infection is essential to provide appropriate treatment for patients and to limit the spread of virus. Rapid global spread of SARS-CoV-2, the causative virus of COVID-19 disease, has led to over 12 million confirmed infections and >500,000 reported deaths worldwide. ![]()
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