Human IFN beta kit HTRF®

Name: Human IFN beta kit
Brand: Cisbio
SKU: 42937
Availability: InStock

Associated products

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

No-wash
Low sample consumption
Under 2h bench time

Download the protocols

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

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Overview

Type I interferons, interferon alpha (IFN-α) and interferon beta (IFN-β), play essential roles in the innate immune response to viral infections. IFN-β is produced by fibroblasts, macrophages, and dendritic cells, as well as endothelial cells. The expression of the IFN-β gene is controlled by transcription factors like IRF-3, c-Jun/ATF-2, and NF-kappaB downstream of Pattern Recognition Receptors, such as Toll-like receptors. In particular, IFN-β is the primary functional readout of cGas-cGAMP-STING pathway activation. Once secreted, IFN-β binds and activates the IFNAR1/IFNAR2 receptor complex, which relies on STAT1/2 transcription factors. Phosphorylated STAT1/2 leads to the induction of anti-viral gene expression.

Assessment of serum samples often requires enhanced sensitivity. In some cases, AlphaLISA assays may have sufficient sensitivity to enable the detection of low levels of analytes in serum or plasma. Check out the Human IFN-β AlphaLISA kit’s analytical performances for more information or learn more about AlphaLISA. When assaying, always follow the recommended protocol and avoid highly haemolyzed samples.

Benefits

SCALABLE FROM 96 TO 1536WELLS
PANEL QUANTIFICATION

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.

To fully understand how to deal with HTRF data processing and also 4PL 1/y² fitting, please visit this page.

Cisbio also worked with Myassays.com to help you in your data analysis. By clicking on this link, you’ll be able to access a free online software to run your IFN beta analysis.

Technical specifications of human IFN beta kit

Sample size	14 µL
Final assay volume	20 µL
Kit components	Lyophilized standard, frozen detection antibodies, activation reagent, buffers & protocol
LOD & LOQ (in Diluent)	7 pg/mL & 19 pg/mL
Range	19 – 4,000 pg/mL
Time to result	3 h at RT
Calibration	NIBSC (00/572) value (IU/ml) = 0.44 x HTRF hIFNβ value (pg/ml)
Species	Human only

Intra and inter assay

Intra assay (n=24)

Sample	Mean [IFN-β] (pg/mL)	CV
1	33	4%
2	321	3%
3	3900	5%
	Mean CV	4%

Inter assay (n=4)

Sample	[IFN-β] (pg/mL)	Mean (delta ratio)	CV
1	62	374	10%
2	329	1628	7%
3	1739	5690	9%
		Mean CV	9%

STING pathway activation by cGAMP triggers hIFNβ release in PBMC

400,000 PBMC were seeded in a 96 well plate under 100 µL of cell culture medium, then stimulated with 50 µL of cGAMP used at 67µM. After an overnight incubation, 14 µL of supernatant were transferred into a white detection plate and human IFNβ was quantified by HTRF.

LPS efficiently induces hIFNβ release in THP1 cells

400,000 THP1cells were seeded in a 96 well plate under 100 µL, then stimulated with 50 µL of LPS used at 20 µg/ml. After 4h30 incubation, 14 µL of supernatant were transferred into a white detection plate, and human IFNβ was quantified by HTRF.

Recombinant THP1 cells expressing constitutively activated STING variant spontaneously releases hIFNβ

400,000 of THP1-WT cells (wild type) and THP1-S154 cells (THP1-Dual™ KI-hSTING-S154 Cells, Invivogen) were seeded in a 96 well plate, under 100 µL of cell culture medium. After 4 h 30 of stimulation, 14 µl of supernatant were transferred into a white detection plate, and human IFNβ was quantified by HTRF.

dsDNA mimetic (CDS agonist) induces hIFNβ secretion

400,000 THP1-S154 cells (THP1-Dual™ KI-hSTING-S154 Cells, Invivogen) were seeded in a 96 well plate under 100 µL of cell culture medium, then treated with dsDNA mimetic (CDS Agonist – LyoVec, Invivogen). After an overnight incubation, 14 µL of supernatant were transferred into a white detection plate, and human IFNβ was quantified by HTRF.

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