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Enzyme-linked immunospot (ELISPOT) technology is a sensitive method that enables visualization of secretory cytokines at the single-cell level. The secretory cytokines are detected using a detection antibody system similar to that used in enzyme-linked immunosorbent assays (ELISA). Cytokines secreted by activated cells are captured by capture antibodies on a membrane and are visualized by detection antibodies in combination with substrates that precipitate at the site of the captured cytokines to form visible spots. The larger the spot and the stronger its staining intensity, the more cytokines are secreted by the corresponding cell over time. However, the appearance of spots can also be strongly influenced by aspects of detection techniques. The ELISPOT assay has the advantage over ELISA in not only measuring cytokine responses at the single-cell level, but also in being able to detect specific cytokine producing cells with higher sensitivity even when the cytokine secreted into the supernatant is below the level detected by ELISA. The highly sensitive ELISPOT technique can detect and analyze cells in very low frequencies within cell populations, down to a small number of cells including single cells. The ELISPOT assay is easy to perform, stable, and allows rapid analysis of large numbers of samples. ELISPOT is not limited to the measurement of cytokines, and is applicable to almost any secreted proteins in single-cell analysis. In addition, the method can readily be qualified and validated.

ELISPOT technology is developed rapidly and has been widely used in immunological research and clinical applications for the determination of antigen-induced or spontaneous specific cytokine responses of various types of immune cells due to microbial pathogens, vaccines, and foreign antigens or autoantigens. The robustness and sensitivity of ELISPOT, together with many new advances in materials, reagents, and protocol steps, have brought this technique into the polyfunctional analysis area. FluoroSPOT is an adaptation of the ELISPOT, which uses fluorescent dyes to quantify cells that secrete analytes. Compared to colorimetric formats, FluoroSPOT is more advantageous in the field of multiplexing and automatic spot detection. The use of fluorescent dyes to measure both analytes simultaneously in a single well allows the identification of subgroups of cells that secrete one or both analytes. In addition, FluoroSPOT requires only half of the cells compared to two ELISPOT assays that analyze one analyte separately. Various analyte combinations allow assess dual or even triple protein secretion patterns at the single-cell level with extremely high sensitivity.

Amerigo Scientific offers highly sensitive, microplate based, and easy-to-operate ELISPOT kits to measure the number of cells secreting cytokines such as interleukin, tumor necrosis factor, or interferon.