HTRF Human IFNγ Detection Kit, 500 Assay Points

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HTRF Human IFNγ Detection Kit, 500 Assay Points

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The HTRF human IFN gamma kit is designed for the quantification of human IFN gamma release in cell supernatant.

For research use only. Not for use in diagnostic procedures. All products to be used in accordance with applicable laws and regulations including without limitation, consumption and disposal requirements under European REACH regulations (EC 1907/2006).

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Feature	Specification
Application	Protein Quantification
Sample Volume	16 µL

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The HTRF human IFN gamma kit is designed for the quantification of human IFN gamma release in cell supernatant.

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Product Variants

Unit Size: 96 Assay Points

Part #:

62HIFNGPET

List Price

USD 652.48

Unit Size: 500 Assay Points

Part #:

62HIFNGPEG

List Price

USD 1,103.00

Unit Size: 10,000 Assay Points

Part #:

62HIFNGPEH

List Price

USD 10,290.00

Product information

Overview

IFN gamma is a pro-inflammatory cytokine secreted by T and NK cells in response to viral infection and or inflammatory conditions. IFN gamma plays a major role in innate and adaptive immunity by promoting T cell differentiation into TH1 and development of Treg, as well as by inhibiting T cell differentiation into TH17. Its anti-tumor properties have been used in treatment of osteoporosis and it is of great interest in immuno-oncology where it improves survival in patients with bladder or ovarian carcinoma.

Assessment of serum samples often requires enhanced sensitivity. In some cases, AlphaLISA assays may have sufficient sensitivity to enable detection of low levels of analytes in serum or plasma.

Specifications

Other Specifications

Application	Protein Quantification
Brand	HTRF
Detection Modality	HTRF
Product Group	Kit
Sample Volume	16 µL
Shipping Conditions	Shipped in Dry Ice
Target Class	Cytokines
Target Species	Human
Technology	TR-FRET
Therapeutic Area	Oncology & Inflammation
Unit Size	500 Assay Points

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Citations

View product citations for 62HIFNGPEG on CiteAb

How it works

Assay principle

Cell supernatant, sample, or standard is dispensed directly into the assay plate for the detection by HTRF® reagents (384-well low-volume white plate or Cisbio low-volume 96-well plate in 20 µl). The antibodies labeled with the HTRF donor and acceptor are pre-mixed and added in a single dispensing step, to further streamline the assay procedure. The assay can be run up to a 1536-well format by simply resizing each addition volume proportionally.

Assay data analysis

The 4 Parameter Logistic (4PL) curve is commonly recommended for fitting an ELISA standard curve. This regression enables the accurate measurement of an unknown sample across a wider range of concentrations than linear analysis, making it ideally suited to the analysis of biological systems like cytokine releases.

Assay details

Technical specifications of human IFN gamma kit

Sample size	16 µL
Final assay volume	20 µL
Kit components	Lyophilized standard, frozen detection antibodies, buffers &protocol.
LOD &LOQ (in Diluent)	14 pg/mL &21 pg/mL
Range	21 – 4,000 pg/mL
Time to result	Overnight at RT
Calibration	NIBSC (82/587) value (IU/mL) = 0,019 x HTRF hIFN? value (pg/mL)
Species	Human only

Analytical performance

Intra and inter assay

Intra-assay (n=24)

Sample	Mean [IFNg] (pg/mL)	CV
1	209	9%
2	835	11%
3	4367	7%
	Mean CV	9%

Each of the 3 samples was measured 24 times, and % CV was calculated for each sample.

Inter-assay (n=4)

Sample	[IFNg] (pg/mL)	Mean (delta R)	CV
1	98	178	14%
2	432	788	9%
3	1905	3170	6%
		Mean CV	9.7%

Each of the samples was measured in 4 different experiments, and % CV was calculated for each sample.

Dilutional linearity

The excellent % of recovery obtained from these experiments show the good linearity of the assay.

Sample	Dilution factor	[IFNg] expected (pg/mL)	[IFNg] detected (pg/mL)	Recovery
1	1	-	150	-
	2.1	72.6	66	91%
	4.3	35	30	86%
			Mean	88%
2	1	-	1953	-
	2.1	942	959	102%
	4.3	454	450	99%
			Mean	100%

Spike and recovery

Sample	[IFNg] added (pg/mL)	[IFNg] expected (pg/mL)	[IFNg] detected (pg/mL)	Recovery
1	2,000	2,421	2,086	86%
2	2,000	3,360	3,363	100%

The same amount of recombinant cytokine was added to 2 different serum samples, and the set of responses obtained from a standard curve was compared to the calculated expected values. The ~ 100% of recovery observed validates the sample matrix used for this assay.

Assay validation

IFNg secretion in PBMCs stimulated with PMA and Ionomycin

PBMC plated at 50, 100, 200 and 400 kcells/well were stimulated for 3h with increasing concentrations of PMA (0, 1, 50 ng/mL) added to a 500 ng/mL Ionomycin solution. 16 µL of supernatants were then transferred into a white detection plate (384, low volume) to be analyzed with the Human IFN? Assay.