Sensitive and portable electrochemical immunoassay for lipoprotein-associated phospholipase A2 using BSA-doped CaCO3 nanospheres to regulate pH readout

摘要

A simple and sensitive electrochemical immunoassay method was developed for the quantitative monitoring of lipoprotein-associated phospholipase A2 (Lp-PLA2) on a low-cost microtiter plate by using a portable handheld pH meter. Bovine serum albumin (BSA)-doped CaCO3 nanospheres were utilized to regulate the pH value of an acidic solution on the basis of classical chemical reactions. To construct such an immunosensing system, the synthesized BSA-CaCO3 nanospheres were initially conjugated covalently to an anti-Lp-PLA2 detection antibody, and then a sandwich-type immunoreaction was carried out on the anti-Lp-PLA2 capture antibody-coated microplate in the presence of the target analyte. Accompanying the sandwiched immunocomplexes, the labeled CaCO3 nanospheres were dissolved in the presence of acid (CaCO3 + 2H+ → Ca2+ + CO2 + H2O), thus resulting in the pH change of the detection solution. Introduction of CaCO3 nanospheres including numerous carbonate ions was expected to enhance the sensitivity of the electrochemical immunoassay. Under optimum conditions, BSA-CaCO3-based immunoassay displayed good pH responses for the determination of target Lp-PLA2 within the dynamic linear range from 0.1 ng mL?1 to 300 ng mL?1. The limit of experimental detection achieved was ～78 pg mL?1 Lp-PLA2 with good coefficients of variation for the inter- and intra-assays. Also, this system gave good anti-interfering capacity toward other biomarkers and enzyme proteins. Importantly, our strategy was applied for the analysis of human serum specimens with satisfactory results in comparison with commercialized Lp-PLA2 ELISA kits.