The Enzyme-Linked ImmunoSpot (ELISpot) assay is a very sensitive immunoassay, allowing the detection of a secreted cytokine at the single cell level. With detection levels that can be as low as one cell in 100 000, the ELISpot is one of the most sensitive cellular assays available. Depending on the substance analyzed, the ELISpot assay is between 20 and 200 times more sensitive than a conventional ELISA. 

n fact, the ELISpot displays a similar sensitivity as a RT-PCR analysis but detects the secreted protein instead of the mRNA. For cytokine analyses this is advantageous since many cytokines are translationally regulated. 

ELISpot analyses are also less impaired by binding proteins and protease activity since the analyte is bound to the capture antibody immediately after secretion. Due to its high sensitivity, the ELISpot has proven particularly useful when studying small populations of active cells such as those regularly found in specific immune responses.

The cytokine ELISpot assay has been widely applied to investigate specific immune responses in various diseases including infections, cancer, allergies and autoimmune diseases.

With Elispot method, active cell or cells that make up the immune response is made visible. Each spot cell is developed from a single cell that is active again. The spots can be counted and from calculating the number of spots in stimulated cultures and control cultures without stimulus, the number of responding cells may be determined.