The Enzyme-linked immunospot (ELISPOT) assay was developed by Czerkinsky in 1983. The ELISPOT assay is based on, and was developed from a modified version of the ELISA immunoassay. It is a sensitive method for monitoring immune responses in humans and animals and a superior tool for accurate quantification of cytokine-secreting T cells.
Nowadays the ELISPOT assay is widely used in different fields of biomedical research including:
Intended use of the ELISPOT assay
The cytokine ELISPOT assay is designed to enumerate cytokine secreting cells in single cell suspensions of lymphoid tissue, CNS tissue, bone marrow or preparations of peripheral blood mononuclear cells (PBMC). The assay has the advantage of detecting only activated/memory T cells and the cytokine release can be detected at the single cell level, allowing direct determination of T cell frequencies. The high sensitivity and easy performance, allowing a direct enumeration of peptide-reactive T cells without prior in vitro expansion, makes the ELISPOT assay eminently well suited to monitor T cell responses
Brief description of the ELISPOT procedure
Cells are incubated for a defined length of time in the wells of the ELISPOT plate precoated with a high-affinity monoclonal antibody to which the cytokine, produced during incubation, will bind. Subsequently, cells are lysed and debris is washed away. Areas in which the cytokine has been captured by the coating antibody are detected with a combination of biotinylated anti-cytokine detector antibodies and enzyme-labeled streptavidin or anti-biotin antibodies. The last step in the assay is the addition of a substrate yielding a colored zone ('spot'), which reveals the site of cytokine secretion.
Eli. Scan is a cost-efficient solution for labs which have only a limited number of plates to measure and no demand for a high throughput system.
It has been designed for the exclusive measurement of punched-out well bottoms from Millipore MultiScreen® plates detached Nunc SilentScreen® membranes and is characterized by a very compact design and a high scanning speed.
The instrument comes as a complete system including scanner unit, PC with Windows 7 Professional®, ELISPOT Analysis Software V5.1, TFT screen, color ink printer and two years warranty. Optionally Eli.Scan can be ordered with a notebook instead of the office PC.
Universal Plate Scanner Eli.Scan Plus offers the complete functionality of the Eli.Scan ELISPOT Scanner
It is able to measure ELISPOT Assays based on detached Nunc® membranes or punched-out well bottoms from Millipore® plates.
In addition it supports the analysis of 6/12/24/48 and 96 well microtiter plates with transparent well bottoms.
The instrument comes as a complete system including scanner unit, PC with Windows 7 Professional®, Eli.Analysis Software V6.0, TFT screen, color ink printer and two years warranty. Optionally it can be ordered with a notebook instead of the office PC.
Eli.Punch ist a tool to easily transfer the well membranes of 96 well Millipore plates to the adhesive foil Eli.Foil. These foils can be read in by our Eli.Scan or Eli.Scan+ instruments. In addition it’s a suitable solution for a space saving storage of your ELISPOT assays
Eli.Foil is a specially designed adhesive foil for punching out the membrane bottoms of 96 well Millipore plates. The foil contains glue only at the locations where the well bottoms will be sticked. This means no more sticking between the bottoms, on the scanner, at your fingers or in your lab journal
Kits are available in different formats (2- or 5-plate format) with different coloring agents (either for silver or enzymatic staining producing "black" and "red" spots, respectively). They can be supplied with transparent polystyrene-bottomed or PVDF membrane-bottomed (PVDF) plates. PB KITS use a silver staining detection method producing “black” spots and PR KITS use an enzymatic detection method producing “red” spots
The antibody pairs have been tested on assay performance with the same secondary reagents and materials as included in the U-CyTech ELISPOT kits. The antibody pairs are supplied in a 10- or 20-plate format (960 or 1920 determinations, respectively) and are delivered with a Technical Data Sheet (TDS). Plates and secondary reagents for ELISPOT performance can be obtained as separate items (see Auxiliary Products ).
The dual-color FluoroSpot assay is a modification of the ELISPOT assay and is designed to detect double cytokine-secreting T cells at the single cell level. The assay is based on the use of fluorescent-labeled conjugates, which can be visualized by fluorescence microscopy or by a fluorescent immunospot image analyzer.
The FluoroSpot assay involves simultaneous staining with both Alexa 488 (green) and R-PE (red) dyes visualizing cells that produce the two cytokines of interest. Cell that release both cytokines become visible as yellow spots. The kits are supplied with PVDF membrane-bottomed (PVDF) plates, which are specially optimized for the FluoroSpot assay.