Cytokine protein arrays.

Abstract

Cytokines play important roles in many aspects of cell physiology and pathology. The simultaneous determination of multiple cytokine-expression levels is receiving much attention in the research community. Multiple cytokine-expression levels can be simultaneously determined using enzyme-linked immunosorbent assay (ELISA)-based protein array technology. In this approach, target proteins are captured by the arrayed capture antibody and then detected in a sandwich ELISA format using a cocktail of biotinylated detection antibodies. The signals are visualized by either horseradish peroxidase (HRP)-conjugated streptavidin and enhanced chemiluminescence, or cy3-conjugated streptavidin and laser scanner. Several key factors and steps are described, including selection of solid supports, selection of suitable antibodies, determination of specificity and sensitivity of cytokine protein arrays, array design, sample preparation, and detailed experimental procedures for macroarray and microarray formats. An account of the successful development and application of cytokine protein arrays is presented.