用纯化的抗体包被微孔板,制成固相载体,往包被抗L-Selectin/CD62L抗体的微孔中依次加入标本或标准品、生物素化的抗L-Selectin/CD62L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终的黄色。颜色的深浅和样品中的L-Selectin/CD62L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。
The microtiter plate provided in this kit has been pre-coated with an antibody specific to L-Selectin. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated polyclonal antibody preparation specific for L-Selectin 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 L-Selectin, 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 L-Selectin in the samples is then determined by comparing the O.D. of the samples to the standard curve.
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