Human TNF-α (Tumor Necrosis Factor Alpha) ELISPOT Kit

This assay kit uses the Sandwich-ELISA principle. The PVDF membrane at the bottom of the micro ELISPOT plate has been re-coated with anti-human TNF-α antibodies. During the experiment, cell suspension and stimulants are added first, and TNF-α secreted by the cells during the incubation process will bind to the coated antibodies. Then wash away the cells, add biotinylated anti-human TNF-α antibodies and alkaline phosphatase conjugated avidin sequentially. The biotinylated anti-human TNF-α antibodies will bind specifically to the TNF-α which has already bounded, then avidin will bind specifically to biotin, together forming the complex. After free components are washed away, add chromogenic substrate (BCIP/NBT), BCIP undergoes hydrolysis under the catalysis of alkaline phosphatase and reacts with NBT, forming insoluble spots with color dark blue to blue-purple. Professional ELISPOT plate reader software can be used to analyze the result, calculating the frequency of human TNF-α secreting cells.