Comparative Analysis of Multiple Immunoassays for Cytokine Profiling in Drug Discovery

Cytokines and their receptors play critical roles in biological processes. Dysfunction or dysregulation of cytokines may cause a variety of pathophysiological conditions. Consequently, cytokine profiling and related technologies are essential for biological studies, disease diagnosis, and drug discovery. In this report, three cytokines, interleukin (IL)-1 beta, IL-6, and tumor necrosis factor alpha (TNF-alpha), from the same sets of samples were analyzed with several commonly used technologies (enzyme-linked immunosorbent assay [ELISA], Luminex, Meso Scale Discovery [MSD], time-resolved fluorescence resonance energy transfer [TR-FRET], cytometric bead array [CBA], AlphaLISA, and FirePlex). Through experimental data analysis, several assay features were compared, including sensitivity, dynamic range, and robustness. Our studies reveal that MSD has the best sensitivity in the low detection limit and the broadest dynamic range, while CBA and Luminex also demonstrate superior performance in the sensitivity and dynamic range. Additional aspects of these technologies, including assay principles, formats, throughputs, robustness, costs, and multiplexing capabilities, were also reviewed and compared. Combining all these features, our comparison highlights MSD as the most sensitive technology, while CBA is the most suitable one for cytokine high-throughput screening with multiplexing capability. Along with perspectives on new technology development in the field, this report aims to help readers understand these technologies and select the proper one for specific applications.