Background: The complex pathology of disease has sparked the development of novel protein expression profiling techniques that require validation in clinical settings. This study focuses on multiplexed analyses of adipocytokines and biomarkers linked to the metabolic syndrome, diabetes, and cardiovascular disease.
Methods: Multiplexed immunoassays using fluorescent microspheres and the Luminex-100 system were performed on plasma from 80 obese patients (40 with the metabolic syndrome) before and after 6-8 weeks of diet-induced weight loss. Leptin, insulin, C-peptide, monocyte chemoattractant protein-1 (MCP-1), eotaxin, interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-alpha), and IL-6 concentrations measured with multiplex panels from 3 different manufacturers were compared with results from commercial ELISAs. Detection limits and between- and within-run imprecision were determined for each analyte. Bland-Altman analysis was used to determine agreement between multiplexed immunoassays and ELISAs.
Results: Correlation between the Luminex multiplexed assays and ELISAs was good for leptin (Linco), insulin (Linco), MCP-1 (Biosource and Upstate), and eotaxin (Biosource) with correlation coefficients of 0.711-0.895; fair for eotaxin (Upstate) and C-peptide (Linco) with correlation coefficients of 0.496-0.582; and poor for TNF-alpha, IL-8, and IL-6 (Linco, Biosource, Upstate, and R&D) with correlation coefficients of -0.107 to 0.318. Within- and between-run imprecision values for the multiplex method were generally <15%. Relative changes in plasma leptin and insulin concentrations after diet-induced weight loss were similar whether assessed by multiplex assay or ELISA.
Conclusion: Although this technology appears useful in clinical research studies, low assay sensitivity and poor correlations with conventional ELISA methods for some analytes with very low plasma concentrations should be considered when using the Luminex platform in clinical studies.