High Sensitive IFN gamma ELISA Assay

Human High Sensitive IFN-gamma ELISA Assay

$765.00

The HHuman High Sensitive IFN-gamma ELISA Assay is to be used for the in-vitro quantitative determination of interferon gamma (IFNγ) in human sera, plasmas, buffered solutions or cell culture media. The assay will recognize both natural and recombinant human IFNγ. The Eagle Biosciences Human IFNγ (Interferon Gamma) ELISA Assay kit is for Research Use Only.

SKU: IFG39-K01 Categories: , , Tag:

Human High Sensitive IFN-gamma ELISA Assay

The Human High Sensitive IFN-gamma ELISA Assay is For Research Use Only

Size: 1×96 wells
Sensitivity: 0.69 pg/ml
Dynamic Range: 0.78 pg/ml – 25 pg/ml
Incubation Time: 3 hours
Sample Type: Serum, Plasma, Cell Culture
Sample Size: 100 µl
Alternative Names: Interferon Gamma, IFN-G

Assay Background

Interferon-gamma (IFN-γ, IFN-g, IFN gamma), also known as type II or immune interferon, is a dimerized soluble cytokine.

Interferon gamma is produced mainly by activated T cells and Natural Killer cells as part of the innate immune response, and by CD4 Th1 and CD8 cytotoxic T lymphocyte (CTL) effector T cells once antigen-specific immunity develops. IFN gamma is also produced by non-cytotoxic innate lymphoid cells (ILC), a family of immune cells first discovered in the early 2010s

IFNg is considered to be a proinflammatory cytokine, that activates macrophages and endothelial cells, it also regulates immune responses by effecting APC, and T cells.

IFN gamma is a cytokine that is critical for innate and adaptive immunity against viral, some bacterial and protozoal infections. IFN gamma is an important activator of macrophages and inducer of Class II major histocompatibility complex (MHC) molecule expression. Aberrant IFNγ expression is associated with a number of autoinflammatory and autoimmune diseases. The importance of IFNγ in the immune system stems in part from its ability to inhibit viral replication directly, and most importantly from its immunostimulatory and immunomodulatory effects.

Different populations of T-cells secrete differing patterns of cytokines that ultimately lead to different immune responses. IFNγ production is a key function of Th1, CD8+ CTLs and also NK cells. It is a cytokine critical for cell mediated immunity against viral and intracellular bacterial infections and is involved in the inflammatory response following secretion via macrophage activation and stimulation of antibody secretion. IFN is the hallmark effector cytokine of Th1 and therefore is an excellent marker for identifying a host response to intracellular pathogens.

IFN is produced during infection by T cells of the cytotoxic/suppressor phenotype (CD8) and by a subtype of helper T cells, the Th1 cells. Th1 cells secrete IL-2, IL-3, TNFα and IFNγ, whereas Th2 cells mainly produce IL-3, IL-4, IL-5, and IL-10, but little or no IFNγ. IFNγ preferentially inhibits the proliferation of Th2 but not Th1 cells, indicating that the presence of IFNγ during an immune response will result in the preferential proliferation of Th1 cells.

In addition, IFNγ has several properties related to immunoregulation. IFNγ is a potent activator of mononuclear phagocytes, and activates macrophages to kill tumor cells by releasing reactive oxygen intermediates and TNFα. IFNγ induces or augments the expression of MHC antigens on macrophages, T and B cells and some tumor cell lines. On T and B cells IFNγ promotes differentiation. It enhances proliferation of activated B cells and can act synergistically with IL-2 to increase immunoglobulin light-chain synthesis.

The role of IFNγ as a disease marker has been demonstrated for a number of different pathological situations including, viral infection, autoimmune disease, transplant rejection, diabetes and allergy.

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Additional Information

Assay Principle

The IFNγ Kit is a solid phase sandwich Enzyme Linked-Immuno- Sorbent Assay (ELISA). A monoclonal antibody specific for IFNγ has been coated onto the wells of the microtiter strips provided. Samples, including standards of known IFNγ concentrations and unknowns are pipetted into these wells. During the first incubation, the IFNγ antigen and a biotinylated monoclonal antibody specific for IFNγ are successively incubated.

After washing, the enzyme (streptavidin-peroxydase) is added. All the unbound enzyme is removed by washing and the first amplification step is performed by adding the Biotine-Tyramine reagent. Under the action of HRP, a biotine polymerisation reaction occurs in the region of the HRP linked to the detection antibody. After washing the second amplification step is performed and the polymerised biotine is revealed by a new streptavidin-HRP step. Finally after washing, the substrate is added. The intensity of this coloured product is directly proportional to the concentration of IFNγ present in the samples.

Assay Procedure

  1. Add 100ml of each standard, sample and zero (Standard Diluent Buffer) in duplicate to appropriate number of wells
  2. Cover with a plastic plate cover and incubate at room temperature (18 to 25°C) with slow shaking for 1 hour
  3. Remove the cover and wash the plate
  4. Add 50ml of diluted biotinylated anti-IFNγ to all wells
  5. Cover with a plastic plate cover and incubate at room temperature (18 to 25°C) with slow shaking for  1 hour
  6. Repeat wash step 3.
  7. Add 100μl of Streptavidin-HRP solution 1 into all wells
  8. Cover with a plastic plate cover and incubate at room temperature (18 to 25°C) with slow shaking for 20 min
  9. Repeat wash step 3.
  10. Add 100ml of diluted Amplifier to all wells
  11. Cover with a plastic plate cover and incubate at room temperature (18 to 25°C) with slow shaking for 15 min
  12. Repeat wash step 3.
  13. Add 100μl of Streptavidin-HRP solution 2 into all wells
  14. Cover with a plastic plate cover and incubate at room temperature (18 to 25°C) with slow shaking for 20 min
  15. Repeat wash step 3.
  16. Add 100μl of ready-to-use TMB Substrate Solution into all wells
  17. Incubate in the dark for 10-20 minutes* at room temperature. Avoid direct exposure to light by wrapping the plate in aluminium foil
  18. Add 100μl of H2SO4:Stop Reagent into all wells

Typical Standard Curve

Human High Sensitive IFN-gamma ELISA Assay

Specificity

Ten specificities were tested with concentrations higher than Interferon Gamma curve concentrations in this Interferon ELISA Assay kit.  No cross reaction was observed for concentrations ranging from 2500 to 78.12 pg/ml for IL-1α, IL-2, IL-8, IL-12p40, TNFα CD95 Fas, TRAIL, ICAM-1, Gp130 and GM-CSF.

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