Mouse IL-1β ELISA Kit

Description

INTENDED USE

This Mouse IL-1β ELISA kit is to be used for the in vitro quantitative determination of mouse interleukin 1 (IL-1β) concentrations in serum, cell culture supernatant, and other biological fluids.  This kit is intended for LABORATORY RESEARCH USE ONLY.

INTRODUCTION

Interleukin 1 (IL-1) is first cytokine super-family identified during the early research on cell secreted factors that regulator fever and inflammatory reaction.  IL-1 is also known as lymphocyte activating factor, endogenous pyrogen, catabolin, hemopoietin-1, melanoma growth inhibition factor, and osteoclast activating factor.  The properties and biological activities of IL-1 have been extensively reviewed.  The two most studied members in the family are Interleukin 1α (IL-1α) and Interleukin 1β (IL-1β).  IL-1α (also known as IL-1F1) and IL-1β (also known as IL-1F2) are distinct gene products, but they recognize the same receptors and have similar biological properties.   A natural antagonist of IL-1α and IL-1β called IL-1Ra was found to compete with IL-1α and IL-1β for membrane receptor binding to down-regulate the activity of IL-1α and IL-1β.   

IL-1β is primarily produced by activated macrophage as a proprotein, which is cleaved by caspase I to become active.  Like IL-1α, IL-1β is an important mediator of innate immunity.  IL-1β stimulates lymphocyte proliferation and the production of TNF-α, IL-6 and IL-8.   IL-1β and TNF-α can synergically act at NFkB and mitogen kinases including JNKs and p38 MARKs, thus leading to the selective expression of genes for immunity and inflammation.

PRINCIPLE OF THE ASSAY

This mouse IL-1β enzyme-linked immunosorbent assay (ELISA) applies a technique called a quantitative sandwich immunoassay.  The microtiter plate provided in this kit has been pre-coated with a monoclonal antibody specific for mouse IL-1β.  Standards or samples are then added to the appropriate microtiter plate wells and incubated.  Mouse IL-1β, if present, will bind and become immobilized by the antibody pre-coated on the wells.  A biotin-conjugated antibody preparation specific for mouse IL-1β is added to each well and incubated.  The biotin labelled antibody will attach to the wells by binding to mouse IL-1β present in the standards/samples.  After plate washing, other proteins, components and unattached biotin labelled antibody are removed.  Then, avidin-horseradish peroxidase (HRP) conjugate is added to each well.  Avidin has a very high affinity for biotin, thus, it links the tracer (HRP) sturdily to the biotin conjugated antibody.  The wells are thoroughly washed to remove all unbound avidin-HRPconjugate and aTMB(3,3’5,5′ tetramethyl-benzidine) substrate solution is added to each well.  The enzyme (HRP) and substrate are allowed to react over a short incubation period.  Only the wells that contain mouse IL-1β will exhibit a change in colour.  The extent of colour change is proportional to the quantity of mouse IL-1β present in the standards/samples.  The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the colour change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm.

In order to measure the concentration of mouse IL- IL-1β in the samples, this kit contains two calibration diluents (Calibrator Diluent I for serum/plasma testing and Calibrator Diluent II for cell culture supernatant/ urine testing).  According to the testing system, the provided standard is diluted (2-fold) with the appropriate Calibrator Diluent and assayed at the same time as the samples.  This allows the operator to produce a standard curve of Optical Density (O.D.) versus IL-1β concentration (pg/mL).  The concentration of IL-1β in the samples is then determined by comparing the O.D. of the samples to the standard curve. 

This IL-1b ELISA is a 3.5-hour solid-phase immunoassay readily applicable to measure IL-1b  levels in serum, cell culture supernatant, and other biological fluids in the range of 0 to 500pg/mL.

CITATIONS  

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2. Hu T, Xu H, Wang C, Qin H, An Z.Magnesium enhances the chondrogenic differentiation of mesenchymal stem cells by inhibiting activated macrophage-induced inflammation. Sci Rep 2018; 1(8):3406.

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