ELISA for interleukin-8 and its primary application

Abstract: To develop a high sensitive and high specific ELISA for IL-8, the high effect antibody was obtained by immunizing rabbits with recombinant IL-8 more times. The purified anti-IL-8 IgG was labeled with biotin and then specially linked to avidin which was labeled with HRP. The purified antibody was used to coat 96 hole board as capture antibody. After adding different concentration of IL-8 standards, reaction was kept for 1.5 hours. Then biotin-IgG, avidin-HRP were added in turn to the reaction system. IL-8 levels were determined by the color, and standard curve was obtained. The IL-8 determine range of this ELISA was 0.2-10.8 ng/mL. CVs within batch and between batch were less than 8% and 10%, respectively. The plasma IL-8 tested in normal persons (n=18) was 0.15 ± 0.06 ng/mL. But in their serum, IL-8 level was tested to be 0.54 ± 0.35 ng/mL by RIA and 0.63 ± 0.59 ng/mL by ELISA. The two methods have obvious correlation in IL-8 values (r=0.819, t=5.89). It was discovered that in cell culture U937 cell stimulated by LPS for 24 or 48 hours produced more IL-8, but U937 cell stimulated by TNF-α for 24 or 48 hours had not produced more IL-8 compared to control. This is one of simple, sensitive and specific method for measurement of IL-8, it can be used to measure IL-8 level in human plasma or serum and the supernatant of cell culture. (authors)