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 one-pot 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.