Description
Human Immunoglobulin G3 (IgG3) ELISpot Antibody Pair Kit is intended for those who want flexibility of setting up their own ELISpot assay. This kit enables detection and enumeration of cells secreting human IgG3 in a peripheral blood mononuclear cell (PBMC) suspension. ELISpot is commonly used for evaluating immune responses, such as determining frequency and isotype or subclass of circulating antigen-specific memory B cells or antibody-secreting cells (ASCs). Together with IgG1, soluble IgG3 has been associated with immune responses to viral infections. ELISpot is highly sensitive and can detect secreted analytes at a single-cell level. This assay is used in research of infectious diseases and vaccines.
This assay can quantify antigen-specific and/or total IgG3-secreting cells. This assay is a modified version of the sandwich ELISA method, in which cells are added to ELISpot plates pre-coated with an antigen of choice or capture antibodies specific for human IgG. Secreted IgG3 is then captured throughout the culture period, before the cells are washed away. Captured IgG3 is detected by the addition of a subclass-specific biotinylated detection antibody, followed by SA-ALP, which binds to the biotinylated antibody. Addition of the chromogenic enzyme substrate 5-bromo-4-chloro-3-indolyl-1-phosphate (BCIP)/nitroblue tetrazolium (NBT) results in the formation of colored spots. The number, size, and intensity of these spots are directly proportional to the frequency of antigen-specific or total IgG3-secreting cells in the sample and amount of analyte secreted.
NOTE: This kit includes sufficient reagents for 4 x 96-well ELISpot plates. ELISpot plates (e.g. Millipore Sigma Catalog #MSIPS4W10 / MAIPSWU10) and BCIP/NBT Substrate (Catalog #200-1016) are required for use with this kit and are available for purchase separately. For detecting antigen-specific immune responses, an antigen of choice is also required.
This kit is supplied in two boxes (Catalog #200-1021/200-1022).
