Description
Introduction: Peroxisome proliferators are non genotoxic carcinogens which are purported to exert their effect on cells through their interaction with members of the nuclear hormone receptor family termed peroxisome proliferator activated receptors (PPARs). Nuclear hormone receptors are ligand dependent intracellular proteins that stimulate transcription of specific genes by binding to specific DNA sequences following activation by the appropriate ligand. Studies indicate that PPARs are activated by peroxisome proliferators such as clofibric acid, nafenopin, and WY14,643, as well as by some fatty acids. It has also been shown that PPARs can induce transcription of acyl coenzyme A oxidase & cytochrome P450 (CYP450) A6 through interaction with specific response elements. The PPAR gamma 2 isoform appears to be induced very early in the differentiation of several cultured adipocyte cell lines, and has been suggested to be a dominant regulator of the murine P2 (aP2) gene which encodes an intracellular lipid binding protein which is expressed only in adipose cells. PPAR gamma 2, like several other nuclear hormone receptors, heterodimerizes with RXR alpha. Defects in PPAR gamma can lead to type 2 insulin-resistant diabetes and hyptertension and could play a role in the genetic predisposition to obesity.
Principle of the Assay: The microtiter plate provided in this kit has been pre-coated with an antibody specific to PPAR-gamma. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific for PPAR-gamma and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB (3,3',5,5' tetramethyl-benzidine) substrate solution is added to each well. Only those wells that contain PPAR-gamma, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm +/- 2 nm. The concentration of PPAR-gamma in the samples is then determined by comparing the O.D. of the samples to the standard curve