Spectrum-Resolved Triplex-Color Electrochemiluminescence Multiplexing Immunoassay with Highly-Passivated Nanocrystals as Tags

Nanocrystals (NCs) were extensively employed in optical-multiplexing with their size-dependent and narrow waveband photoluminescence (PL). Herein, to achieve NCs-based novel optical-multiplexing strategy with improved sensitivity, a spectrum-resolved triplex-color electrochemiluminescence (ECL) multiplexing immunoassay (MIA) was proposed for the first time by extensively exploring the color-different monochromatic ECL from dual-stabilizers-capped CdTe and CdSe NCs and the inherent lower background of ECL than PL. As a proof of concept, carcinoembryonic antigen (CEA), prostate specific antigen (PSA), and alpha fetoprotein (AFP) were adopted as model analytes and formed three antigen-NCs pairs at the glassy carbon electrode surface via a one-pot immune-reaction with CdSe(550), CdTe(650), and CdTe(776) NCs as tags, respectively. The spectral ECL responses of three antigen-NCs pairs were simultaneously measured via onepot ECL assay, and the maximum ECL intensity of each tag was directly utilized to determine corresponding antigen without any assistance from either optical filters or signal deconvolution. This ECL-MIA displayed negligible cross-reactivity, high color-selectivity, and excellent sensitivity with limits of detection down to 1, 10, and 0.01 pg/mL for CEA, PSA, and AFP, respectively, which might provide a promising alternative to NCs PL multiplexing.