Description
Introduction: Macrophage Inflammatory Proteins (MIP) belong to the family of chemotactic cytokines known as chemokines. In humans, there are two major forms, MIP-1alpha and MIP-1beta that are now officially named CCL3 and CCL4 respectively. C-C motif chemokine 3 is a protein that in humans is encoded by the CCL3 gene. Macrophage inflammatory protein-1 is a so-called monokine that is involved in the acute inflammatory state in the recruitment and activation of polymorphonuclear leukocytes (Wolpe et al., 1988).
Principle of the Assay: The microtiter plate provided in this kit has been pre-coated with an antibody specific to MIP-1alpha. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated polyclonal antibody preparation specific for MIP-1alpha 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 MIP-1alpha, 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 MIP-1alpha in the samples is then determined by comparing the O.D. of the samples to the standard curve