Spectrum-Resolved Dual-Color Electrochemiluminescence Immunoassay for Simultaneous Detection of Two Targets with Nanocrystals as Tags.

A spectrum-resolved dual-color electrochemiluminescence (ECL) immunoassay was designed and implemented to simultaneously detect carcinoembryonic antigen (CEA) and alpha fetoprotein (AFP) with CdTe (λmax = 776 nm) and CdSe (λmax = 550 nm) nanocrystals (NCs) as ECL tags. The CdTe and CdSe NCs were labeled with respective probe antibodies (Ab2) of CEA and AFP, respectively, and then immobilized onto the working electrode surface via sandwich-type immunoreactions. Both CdTe and CdSe NCs within the NCs immunocomplexes can be electrochemically reduced and simultaneously give off monochromatic ECL emissions in the near-infrared and greenish regions, respectively, when (NH4)2S2O8 was used as a cathodic ECL coreactant. The ECL spectra of the two surface-confined NCs were well separated and had no cross energy-transfer interactions, which made the dual-color immunoassay highly selective and sensitive toward respective target analytes. With the proposed ECL biosensor, CEA and AFP were simultaneously detected and quantified with an extremely low detection limit of 1 pg/mL for CEA and 10 fg/mL for AFP, respectively. This work demonstrated the probability of performing multianalyte assays via a spectrum-resolved ECL strategy with improved sensitivity and signal-to-noise ratio as compared to NCs-based fluorescent multianalyte assays.