Microchip fluorescence-enhanced immunoaasay for simultaneous quantification of multiple tumor markers

A microchip fluorescence-enhanced immunoassay method was developed for simultaneous detection of carcinoma antigen 125 (CA125) and carbohydrate antigen 15-3 (CA15-3). In this method, CA125 and CA15-3 react with excess amount of fluorescein isothiocyanate (FITC)-labeled monoclonal antibodies (Ab*) of CA125 and CA15-3 to form CA125-Ab125* and CA15-3-Ab15-3* complexes. Microchip electrophoresis (MCE) separation of free Ab125*, Ab15-3*, and CA125-Ab125*, CA15-3-Ab15-3* complexes were then performed. The separated species were sensitively detected by laser-induced fluorescence detection (LIF). CA125 and CA15-3 were quantified simultaneously by measuring the fluorescence intensity of CA125-Ab125* and CA15-3-Ab15-3* complexes, respectively. Under the optimum conditions, the limits of detection were 0.23 U/mL for CA125 and 0.09 U/mL for CA15-3. The present MCE-LIF method was applied to the determination of CA125 and CA15-3 in serum from healthy subjects and cancer patients. The levels of CA125 and CA15-3 in these sera samples were found to be in the ranges of 15.6–36.1 U/mL and 13.8–28.4 U/mL for healthy subjects, and 192.5–368.3 U/mL and 63.3–198.4 U/mL for cancer patients.