Human IFN-gamma Quantikine QuicKit ELISA

Name: Human IFN-gamma Quantikine QuicKit ELISA
Brand: R&D Systems
SKU: QK285
Rating: 4 (1 reviews)

Catalog #: QK285
2 Citations 1 Review Datasheet / COA / SDS

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QK285

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QC290: Human IFN-gamma Quantikine QuicKit Control Group 290

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Human IFN-gamma QuicKit ELISA Spiked Recovery Competitor Comparison

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Summary Sample Values Precision Recovery Linearity Data Examples Product Datasheets Preparation and Storage Background Related Research Areas

Human IFN-gamma Quantikine QuicKit ELISA Summary

Assay Length

80 minutes

Sample Type & Volume Required Per Well

Cell Culture Supernates (50 uL), Serum (50 uL), EDTA Plasma (50 uL), Heparin Plasma (50 uL)

Sensitivity

2.56 pg/mL

Assay Range

31.3 - 2,000 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma)

Cross-reactivity

< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.

Interference

No significant interference observed with available related molecules.

Sample Values

Serum/Plasma - Ten serum and plasma samples from apparently healthy volunteers were evaluated for the presence of human IFN-gamma in this assay. No medical histories were available for the donors used in this study. All samples measured less than the lowest standard, 31.3 pg/mL.

Cell Culture Supernates:

Peripheral blood mononuclear cells (multiple donors) (seeded at 1 x 106 /mL) were cultured in RPMI 1640 supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin sulfate. Cells were left untreated or treated with 10 μg/mL of PHA for 5 days. Aliquots of the cell culture supernates were removed and assayed for levels of human IFN-gamma.

CD4+ T cells were isolated from human PBMCs (single donor) using the MagCellect™ Human CD4+ T cell Isolation Kit (R&D Systems, Catalog # MAGH102). CD4+ T cells were then seeded at 5 x 105 /mL and cultured in ExCellerate Human T Cell Expansion Media, Xeno-Free (R&D Systems, Catalog # CCM030). T cells were left untreated or treated for 5 days. Treated T cell media was supplemented with 10 ng/mL GMP recombinant human (rh) IL-7 (R&D Systems, Catalog # 207-GMP) and 10 ng/mL GMP rhIL-15 (R&D Systems, Catalog # 247-GMP). TCR stimulation was mediated using either (1) immobilized Human CD3 epsilon Antibody (R&D Systems, Catalog # MAB100, coated at 1 μg/mL) with 5 μg/mL soluble Human CD28 Antibody (R&D Systems, Catalog # MAB342) or (2) 25 μL Cloudz CD3/28 particles (Cloudz™ T Cell Activation Kit-CD3/CD28, R&D Systems, Catalog # CLD001) per mL of culture media. Aliquots of the cell culture supernates were removed and assayed for levels of human IFN-gamma.

Condition	(pg/mL)
Unstimulated	ND
Stimulated PBMCs	111,232
Unstimulated CD4+ T Cell	ND
CD4+ T Cell CD3/CD28	52,293
CD4+ T Cell Cloudz	191,034

ND=Non-detectable

Product Summary

The Quantikine QuicKit Human IFN-gamma Immunoassay is a one step, 80-minute solid phase ELISA designed to measure human IFN-gamma levels in cell culture supernates, serum, and plasma. It contains HEK293-expressed recombinant human IFN-gamma and antibodies raised against the recombinant protein. Results obtained using natural human IFN-gamma showed linear curves that were parallel to the standard curves obtained using the QuicKit standards. These results indicate that this kit can be used to determine relative mass values for natural human IFN-gamma.

Precision

Intra-Assay Precision (Precision within an assay) Two samples of known concentration were tested twenty times on one plate to assess intra-assay precision

Inter-Assay Precision (Precision between assays) Two samples of known concentration were tested in ten separate assays to assess inter-assay precision. Assays were performed by at least three technicians

Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma

	Intra-Assay Precision		Inter-Assay Precision
Sample	1	2	1	2
n	20	20	10	10
Mean (pg/mL)	198	1156	211	1233
Standard Deviation	3.76	20.2	20.4	56.9
CV%	1.9	1.8	9.6	4.6

Recovery

The recovery of human IFN-gamma spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type	Average % Recovery	Range %
Cell Culture Media (n=5)	92	83-102
EDTA Plasma (n=4)	107	88-132
Serum (n=4)	93	74-114

Linearity

To assess the linearity of the assay, samples containing and/or spiked with high concentrations of human IFN-gamma were diluted with calibrator diluent to produce samples with values within the dynamic range of the assay. Samples were diluted prior to assay.

Scientific Data

ELISA

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Human IFN-gamma QuicKit ELISA Spiked Recovery Competitor Comparison IFN-gamma is spiked at three known concentrations throughout the range of the assay and run to measure response of the spiked sample matrix. Serum recovery is 115% compared to 144% for the top competitor. Plasma recovery is 120% compared to 135% for the top competitor. Cell Culture Supernates recovery is 118% compared to 79% for the top competitor. In spike and recovery experiments, natural samples are spiked with the recombinant target analyte of interest to identify interference caused by sample matrices.

ELISA

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Human IFN-gamma QuicKit ELISA Spiked Linearity Competitor Comparison IFN-gamma is spiked at high concentration in various matrices and diluted with appropriate Calibrator Diluent to produce samples with values within the dynamic range of the assay. The linearity is between 99%-126% compared to 71%-85% for the top competitor.

N/A

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Human IFN-gamma Standard Curve

Product Datasheets

Product Datasheet

COA

SDS

Preparation and Storage

Shipping

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.

Storage

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: IFN-gamma

IFN-gamma (Interferon-gamma) is the prototype proinflammatory cytokine and is produced by a variety of immune cells under inflammatory conditions, notably by T cells and NK cells. It plays a key role in host defense by promoting the development and activation of Th1 cells, chemoattraction and activation of monocytes and macrophages, upregulation of antigen presentation molecules, and immunoglobulin class switching in B cells. It also exhibits antiviral, antiproliferative, and apoptotic effects. In addition, IFN-gamma functions as an anti-inflammatory mediator by promoting the development of regulatory T cells and inhibiting Th17 cell differentiation. IFN-gamma dimers signal through a receptor complex of two IFN-gamma R1 and two IFN-gamma R2 subunits.

Long Name:

Interferon gamma

Entrez Gene IDs:

3458 (Human); 15978 (Mouse); 25712 (Rat); 396991 (Porcine); 281237 (Bovine); 403801 (Canine); 493965 (Feline)

Alternate Names:

IFG; IFI; IFNG; IFNgamma; IFN-gamma; Immune interferon; interferon gamma; interferon, gamma

Related Research Areas

Citations for Human IFN-gamma Quantikine QuicKit ELISA

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

2 Citations: Showing 1 - 2
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