U-CyTech biosciences is a registered Dutch company engaged in fundamental and applied research in the field of cytokine biology. U-CyTech develops, produces and sells a wide range of innovative cytokine ELISA, ELISPOT and Fluorospot assays for human, monkey and rodent use. The reagents for these systems are available in two different formats: complete kits and matched antibody pairs.
With a wide range of "in house" developed, highly specific monoclonal and polyclonal antibodies directed against a large number of cytokines from various species, U-CyTech has high quality ELISA and ELISPOT kits.
The validated compatibility of these antibodies with certain high-quality auxiliary reagents ensures trouble-free operation and optimal results. Additionally, the FluoroSpot assay is designed to identify individual cells that simultaneously secrete two different cytokines. Dual cytokine secreting cells are visualized by using two fluorescent - instead of enzyme - labeled detection antibodies.
U-CyTech has developed various ELISA kits for the detection of cytokines in cell culture supernatant, plasma or serum. U-CyTech's "sandwich" type ELISA is a simple and sensitive assay for the determination of cytokine levels in biological fluids. The assay is performed in antibody-coated microtiter 96-wells plate and serial dilutions with unknown concentrations of cytokines are added into the wells. The bound cytokine is then associated with a biotinylated detection antibody and an enzyme-labeled streptavidin polymer. Subsequently, a chromogenic substrate is introduced that yields a colored product that can be determined spectrophotometrically. If performing a quantitative ELISA, U-CyTech's ELISA kits provide an appropriate cytokine standard to determine the actual concentration of cytokines in samples.
U-CyTech offers a wide range of ELISA kits for the detection of human and monkey cytokines or granzymes in cell culture supernatant, plasma or serum: e.g. IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-10, IL-17A, GM-CSF, Granzyme B, Perforin, TNF-α.
The monkey ELISA kits have been validated for several Old World monkey species including rhesus macaques, cynomolgus monkeys, baboon, barbary macaques, lion-tailed macaques, pig-tailed macaques, Sulawesi-crested macaques and Japanese macaques. U-CyTech has also developed ELISA kits for the detection of cytokines from the Marmoset (New World monkey species), e.g. IFN-γ, TNF-α and IL-17A.
In addition, U-CyTech offers several ELISA kits for the detection of rat (IFN-γ, IL-4, TNF-α) and mouse (IFN-γ, IL-2, IL-4, IL-5, IL-6, IL-10, TNF-α) cytokines in cell culture supernatant, plasma or serum.
T cell ELISPOT assay
The cytokine ELISPOT assay is designed to enumerate cytokine secreting cells in single cell suspensions of lymphoid tissue, peripheral blood mononuclear cells (PBMC), CNS tissue or bone marrow. The assay has the advantage that only activated/memory T cells are detected, 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 ideally suited to monitor T cell responses.
Cells are incubated in the wells of a 96-well ELISPOT plate precoated with a high-affinity antibody, which captures the secreted cytokine or granzyme. Subsequently, cells are lysed and debris is washed away. Then, biotinylated anti-cytokine detection antibodies are added that bind to the captured product. The addition of enzyme-labeled streptavidin polymer and a substrate, results in a colored zone ("spot"). Colored spots reveal the site of cytokine secretion and represent individual secreting T cells. These cells are identified and counted using an automated ELISPOT reader.
The T cell ELISPOT kits that are offered by U-CyTech are kits for the detection of human and monkey cytokine or granzymes producing cells, including kits, e.g. for IFN-γ, IL-1β, IL-2, IL-4, IL-17A, IL-21, GM-CSF, Granzyme B and TNF-α.
The antibodies in the monkey kits have been validated for detecting cytokine producing cells of various Old World monkeys including Rhesus macaques, cynomolgos monkeys, baboons, barbary macaques, lion-tailed macaques, pig-tailed macaques, Sulawesi macaques and Japanese macaques. Our ELISPOT kits are also available for Marmoset (New World Monkey species), like IFN-γ, IL-12/23p40, IL-13, IL-17A and TNF-α ELISPOT.
Additionally, U-CyTech offers various rat (IFN-γ, IL-4) and mouse (IFN-γ, IL-2, IL-4, IL-5, IL-10, TNF-α) cytokine ELISPOT kits.
B cell ELISPOT assay
The B cell ELISPOT assay is designed for the identification and enumeration of individual antibody-secreting B cells in single cell suspensions of peripheral blood mononuclear cells (PBMC) from humans or monkeys. The technique is based on the same solid-phase immuno-enzymatic principle as the T cell ELISPOT. The first step in the assay is the expansion of memory B cells by polyclonal or antigenic stimulation. Subsequently, expanded B cells are enumerated in the ELISPOT 96-well plate coated with anti-immunoglobulines or an antigen of interest. Thereafter, spot forming cells are detected by labeled antibodies directed against an immunoglobulin class of choice.
Each kit contains a reagent for the stimulation of memory B cells, coating antibodies for the determination of the total number of antibody secreting cells, labeled antibodies for the detection of the immunoglobulin class of choice and coloring agents.
Additionally, U-CyTech offers several mouse (IgG, IgG1, IgG2a, IgG2b) B cell ELISPOT kits. Other B cell ELISPOT kits are under development.
The dual-color FluoroSpot assay is a modification of the ELISPOT assay and developed for the detection of two cytokines released by a single T cell. The assay is based on the use of antibodies labeled with different fluorophores, allowing the detection of T cells producing two pre-selected cytokines. The Fluorospot assay is particularly useful for the analysis of T cell subpopulations with distinct cytokine profiles. Kits are supplied with PVDF membrane-bottomed (PVDF) plates, which are specially optimized for the FluoroSpot assay.
U-CyTech offers a wide range of human and monkey cytokine FluoroSpot kits including kits for the combinatorial detection of IFN-γ and a secondary cytokine (e.g. IL-2, IL-10). Fluorospot kits for detection of multiple cytokines are under development.