This HTRF kit is designed for the quantification of human IL1 beta release in cell supernatant.

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  • No-wash
  • Low sample consumption
  • Under 2h bench time

This HTRF kit is designed for the quantification of human IL1 beta release in cell supernatant.

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In stock

Overview

IL1 beta, also known as leukocytic pyrogen or mononuclear cell factor, is a pro-inflammatory cytokine of the IL1 family. IL1 beta is considered to be a major endogenous pyrogen and induces prostaglandin synthesis, T cell activation, B cell activation, and subsequent antibody production. It is also a known promoter of T cell differentiation into Th17.

Benefits

  • VALIDATED ON THP1 AND PBMC
  • 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 IL1 beta analysis.

Technical specifications of human IL1 beta kit

Sample size 16 µL
Final assay volume 20 µL
Kit components Lyophilized standard, frozen detection antibodies, buffers &protocol.
LOD &LOQ (in Diluent) 5 pg/mL &39.4 pg/mL
Range 39.4 – 6,500 pg/mL
Time to result Overnight at RT
Calibration NIBSC (86/680) value (IU/mL) = 0,1 x HTRF hIL1ß value (pg/mL)
Species Human only

Intra and inter assay

Intra-assay (n=24)

Sample Mean [IL1ß] (pg/mL) CV
1 65 18%
2 705 9%
3 5030 4%
Mean CV 10%

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

Inter-assay (n=4)

Sample [IL1ß] (pg/mL) Mean (delta R) CV
1 159 378 15%
2 702 1742 13%
3 3095 7736 12%
Mean CV 13%

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

Dilutional linearity

Sample
Dilution factor[IL1β] expected (pg/mL)[IL1β] detected (pg/mL)Recovery
11-5486-
4.31276120294%
10.751347993%
Mean93%
21---
4.340837893%
8.919719499%
18.5959598%
Mean97%

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

Spike and recovery

Sample
[IL1β] added (pg/mL)[IL1β] expected (pg/mL)[IL1β] detected (pg/mL)Recovery
1325050575323105%
2325073267488102%



Mean Recovery104%

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.

IL1ß secretion in THP1 cells stimulated with LPS & PMA

THP1 cells plated at 50 and 100 kcells/well were stimulated for 18 h with PMA and LPS respectively used at 50 ng/mL and 1 µg/mL. 16 µL of supernatants were then transferred into a white detection plate (384 low volume) to be analyzed with the Human IL1ß Assay.

IL1ß secretion in PBMC stimulated with LPS

PBMC plated at 50, 100, 200 and 400 kcells/well were stimulated for 18 h with increasing concentrations of LPS ranging from 0.02 to 2 µg/mL. 16 µL of supernatants were then transferred into a white detection plate (384 low volume) to be analyzed by the Human IL1ß Assay Kit.

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