Sandwich Elisa for Detection of Picogram Quantities of Interleukin-8

Abstract

A sensitive sandwich enzyme-linked immunosorbent assay (ELISA) was developed for interleukin-8 (IL-8), a neutrophil chemoattractant and activator. A polyclonal antibody to recombinant human IL-8 was raised in rabbits, and the IgG was isolated from the antisera using a protein A column. Native and biotinylated forms of this antibody served as the capture antibody and developing antibody for the ELISA, respectively, and avidin-conjugated horse radish peroxidase provided the means for enzymatic color development. The lower limit of sensitivity for the assay was found to be 84 ± 20 pg/ml IL-8 (mean ± SD for 10 determinations). An inter-assay variability of 15-29% and an intra-assay variability of 12% were observed. The assay was able to detect IL-8 when the samples were prepared in either normal saline, RPMI, or human plasma. The development of this rapid, sensitive assay should provide a means to more fully evaluate the role of this cytokine in diverse disease states.