Mouse IL-17A ELISA Kit

INTENDED USE 

This mouse Interleukin-17A ELISA Kit is designed for the in vitro quantitative determination of mouse interleukin-17A (IL-17A) concentrations in cell culture supernatant and mouse serum.  This kit is intended for LABORATORY RESEARCH ONLY. 

INTRODUCTION 

Mouse Interleukin-17A (mIL-17A), also know as CTLA-8, is a secreted, homodimeric glycoprotein linked by disulfide link with a molecular mass of about 35 KD. The cytokine is the proto-type of a newly discovered pro-inflammatory cytokine family which consists of IL-17A, IL-17B, IL-17C, IL-17D, IL-17E (IL-25) and IL-17F. IL-17A is mainly produced by a group of CD4 T helper cells termed Th17 cells.  In addition to Th17 cells, CD8 T helper cells, natural killer cells and granulocytes, can also secret the cytokine after proper stimulation.  IL17A exhibits structural homology and cross-species bioactivity between human and mouse cells.    

IL-17A acts through its receptor IL-17R.  It was found to stimulate the expression of pro-inflammatory cytokines including IL-6, MCP-1, G-CSF,GM-CSF,IL-1alpha and TNF-alpha.

Most of those cytokines plays a role in the proliferation, maturation and chemotaxis of neutrophils.  Elevated levels of IL-17A have been found to implicate with autoimmune diseases, airway inflammation, allograft rejection, inflammatory bowel disease, psoriasis, cancer, vascular inflammation and development of atherosis.  

This mouse IL-17A ELISA is a 3.5-hour solid phase immunoassay readily applicable to measure mouse IL-17A levels in cell culture supernatant and mouse serum in the range of 0 to 1000 pg/mL.  It showed no cross reactivity with other cytokines tested.  This mouse IL-17A ELISA is expected to be effectively used for further investigations into the relationship between IL-17A and the various conditions mentioned. 

PRINCIPLE OF THE ASSAY 

This mouse IL-17A 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 to mouse IL-17A.  Standards or samples are then added to the appropriate microtiter plate wells.  A biotin-conjugated antibody preparation specific for mouse IL-17A was added and incubated.  Mouse IL-17A, if present, will bind and become immobilized by the antibody pre-coated on the wells.  The microtiter plate wells are thoroughly washed to remove unbound mouse IL-17A and other components in the sample.  Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated.  Avidin is a tetramer containing four identical subunits that each has a high affinity-binding site for biotin.  The wells are thoroughly washed to remove all unboundHRP-conjugated Avidin, 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 those wells that contain mouse IL-17A, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in colour.  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-17A in the samples this kit includes two calibration diluents (Calibrator Diluent I for serum testing and Calibrator Diluent II for cell culture supernatant 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 mouse IL-17A concentration (pg/mL).  The concentration of IL-17A in the samples is then determined by comparing the O.D. of the samples to the standard curve.

CITATIONS  

1. Wei Y, Dong M, Zhang H, Lv Y, Liu J, Wei K, Luo Q, Sun J, Liu F, Xu F, Dong J. Acupuncture Attenuated Inflammation and Inhibited Th17 and Treg Activity in Experimental Asthma. Evid Based Complement Alternat Med 2015; (2015):340126.