Human Granzyme B ELISpot Kit

Human Granzyme B ELISpot Kit

Assay Genie ELISpot is a highly specific immunoassay for the analysis of Granzyme B production and secretion from T-cells at a single cell level in conditions closely comparable to the in-vivo environment with minimal cell manipulation. This technique is designed to determine the frequency of Granzyme B producing cells under a given stimulation and the comparison of such frequency against a specific treatment or pathological state. Utilising sandwich immuno-enzyme technology, Assay Genie ELISpot assays can detect both secreted Granzyme B (qualitative analysis) and single cells that produce Granzyme B (quantitative analysis). Cell secreted Granzyme B is captured by coated antibodies avoiding diffusion in supernatant, protease degradation or binding on soluble membrane receptors. After cell removal, the captured Granzyme B is revealed by tracer antibodies and appropriate conjugates.

Human Granzyme B ELISpot Kit

The Human Granzyme B ELISPOT Kit is a powerful tool for detecting and quantifying levels of granzyme B in human samples, including serum, plasma, and cell culture supernatants. This kit offers high sensitivity and specificity, allowing for accurate and reproducible results for a variety of research applications.Granzyme B is a key player in the immune response, particularly in the cytotoxic activity of cytotoxic T lymphocytes and natural killer cells.

Its role in inducing apoptosis in target cells makes it a valuable biomarker for studying immune responses in conditions such as infections, autoimmune diseases, and cancer.With the Human Granzyme B ELISPOT Kit, researchers can gain valuable insights into the immune response and the role of granzyme B in various diseases, paving the way for the development of novel therapeutic strategies.

A capture antibody highly specific for Granzyme B is coated to the wells of a PVDF bottomed 96 well microtitre plate either during kit manufacture or in the laboratory. The plate is then blocked to minimise any non-antibody dependent unspecific binding and washed. Cell suspension and stimulant are added and the plate incubated allowing the specific antibodies to bind any Granzyme B produced. Cells are then removed by washing prior to the addition of Biotinylated detection antibodies which bind to the previously captured Granzyme B. Enzyme conjugated streptavidin is then added binding to the detection antibodies. Following incubation and washing, substrate is then applied to the wells resulting in coloured spots which can be quantified using appropriate analysis software or manually using a microscope.

Figure: Schematic representation of the principle of ELISpot. 1) Treat with ethanol, coat with capture antibody. 2) Incubate cells in the presence of stimuli, cytokines, are released, which bind to capture antibodies. 3) Wash plate. Incubate with biotinylated detection antibody. 4) Wash plate. Incubate with streptavidin-alkaline phosphatase conjugated. 5)Incubate with BCIP/NBT. Precipitated product forms, results in blue/purple spots.