*4.1. Human Samples*

Human subjects research was performed according to University of Pittsburgh approved IRB protocols (20030228 and 20040220). Positive and negative control samples used for specificity and sensitivity assays were from residual samples at UPMC clinical laboratories and have been previously described [15]. WHO international standards were from the National Institute for Biological Standards and Control (NIBSC). Serum or plasma was collected and stored at −20 ◦C.

Blood samples obtained in Fall 2020 were from two cohorts. One cohort included 100 samples obtained between 21 September 2020 and 13 October 2020 from healthy adults who were willing to have phlebotomy performed for infectious disease research. All participants self-reported their age, sex, race, ethnicity, residential zip code, history of travel and immunization, as well as history of known COVID-19 disease or contact with COVID-19 confirmed cases. A second cohort of 99 samples were residual specimens obtained from the UPMC Mercy clinical laboratory between 2 November 2020 to 4 November 2020; these specimens were from outpatients undergoing routine bloodwork. Blood samples from February 2021 were all residual specimens obtained from the UPMC Mercy outpatient laboratory between 1 February 2021 and 5 February 2021. Age, sex, race, residential zip code and any history of COVID-19 disease or testing was abstracted from the medical record prior to de-identification for all UPMC Mercy outpatient samples.
