Intended Use：
The kit applies for the detection of mouse IFN-γ at the cellular level by counting IFN-γ-secreting cells.
PRINCIPLE OF THE ASSAY：
The enzyme-linked immunospot (ELISPOT) assay employs a sandwich ELISA-based methodology. A 96-well polyvinylidene difluoride (PVDF) membrane plate, pre-coated with mouse IFN-γ-specific monoclonal antibodies, captures mouse IFN-γ secreted by activated cells. Following cell removal and washing, biotin-conjugated secondary antibodies targeting IFN-γ are added to form antibody-antigen-antibody "sandwich" complexes. Subsequent incubation with streptavidin-alkaline phosphatase (ALP) conjugates enables biotin-mediated binding. Addition of chromogenic substrate culminates in the formation of insoluble spots at the membrane interface, with each spot representing an individual IFN-γ-secreting cell. Spot quantification is achieved by automated ELISPOT analysis or manual counting using a microscope.
