SARS-CoV-2 interferon gamma release assay
The EUROIMMUN SARS-COV-2 interferon gamma release assay (IGRA) (Quan-T-Cell SARS-CoV-2) was used for the assessment of patients’ T-cell response to SARS-CoV-2 antigens through analysis of the production of interferon gamma by patient T cells after exposure to SARS-CoV-2-specific proteins. The assay does not differentiate between vaccine- or infection-induced T-cell responses. The SARS-CoV-2 IGRA is performed in two steps as per the manufacturer’s instructions, and a brief protocol follows. First, patient samples from lithium heparin vacutainers are aliquoted into three separate tubes each. These tubes contain either nothing (‘blank’), general T-cell activating proteins (‘mitogen’), or components of the S1 domain of SARS-CoV-2 (‘SARS-CoV-2 activated’). These samples were incubated at 37°C for 24 hr before being centrifuged and the plasma separated and frozen at –80°C for later analysis. Samples were then batched to be run as a full 96-well plate along with calibrators and controls. Plasma samples were unfrozen and added to an ELISA plate, which was prepared with monoclonal interferon-gamma binding antibodies, along with calibrators and controls. After incubation at RT the plate was washed and biotin-labeled anti-interferon gamma antibody was added to bind the patient interferon gamma bound to the plate. The plate was again incubated before being washed of excess antibody and a streptavidin-bound horseradish peroxidase (HRP) enzyme added, which binds strongly to the biotin-labeled antibodies present. This was again incubated and then washed of excess enzyme before a solution of H