ELISA法定量测定豚鼠血清、血浆或其它相关液体中IL-12含量。

 

本试剂盒应用双抗体夹心酶标免疫分析法测定标本中IL-12水平。用纯化的抗体包被微孔板，制成固相抗体，往包被单抗的微孔中依次加入IL-12抗原、生物素化的抗豚鼠IL-12抗体、HRP标记的亲和素，经过彻底洗涤后用底物TMB显色。TMB在过氧化物酶的催化下转化成蓝色，并在酸的作用下转化成最终的黄色。颜色的深浅和样品中的IL-12呈正相关。用酶标仪在450nm波长下测定吸光度（OD值），计算样品浓度。

 

This immunoassay kit allows for the specific measurement of Guine Pig Interleukin,IL-12 concentrations in cell culture supernates, serum and plasma.

 

Interleukin 12 (IL-12), also known as natural killer cell stimulatory factor (NKSF) or cytotoxic

lymphocyte maturation factor (CLMF), is a pleiotropic cytokine originally identified in the medium of cultured EBV-transformed RPMI-8866 cells . IL-12 is a 75 kDa glycoprotein heterodimer composed of two genetically unrelated subunits linked by a disulfide bond. The smaller subunit (p35) has homology to IL-6 and G-CSF while the larger subunit (p40) demonstrates similarity to the soluble receptor for IL-6, leading to the suggestion that IL-12 might have evolved from a cytokine/soluble receptor complex .

Each subunit of IL-12 apparently arises from a single copy gene. The mRNA transcription of the subunits is closely coordinated, although an excess of the larger subunit has been shown to be produced by B cells in addition to active IL-12 . Expression of p35 is reported to be enhanced by simultaneous expression of p40. IL-12 activity cannot be demonstrated in the absence of either chain . As suggested by their names, p35 has a native molecular weight of 35 kDa while p40 has a native molecular weight of 40 kDa.

IL-12 is produced by macrophages and B lymphocytes and has been shown to have multiple effects on T cells and natural killer (NK) cells . These include inducing production of IFN-gama and TNF by resting and activated T and NK cells, synergizing with other IFN-gama inducers at both the transcriptional and post-transcriptional levels to induce IFN- gama gene expression, enhancing the cytotoxic activity of resting NK and T cells, inducing and synergizing with IL-2 in the generation of lymphokine-activated killer (LAK) cells, acting as a comitogen to stimulate proliferation of resting T cells, and inducing proliferation of activated T and NK cells .

Evidence indicates that IL-12, produced by macrophages in response to infectious agents, is a central mediator of the cell-mediated immune response by its actions on the development, proliferation, and activities of TH1 cells . These activities of IL-12 are antagonized by IL-4 and IL-10, factors associated with the development of uncommitted T helper cells into TH2 cells and mediation of the humoral immune response .

 

This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for IL-12 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any IL-12 present is bound by the immobilized antibody. An enzyme-linked polyclonal antibody specific for IL-12 is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of IL-12 bound in the initial step. The color development is stopped and the intensity of the color is measured.