Fluorescence ELISA based on glucose oxidase-mediated fluorescence quenching of quantum dots for highly sensitive detection of Hepatitis B

Abstract

Herein, we present a novel sandwich fluorescence enzyme linked immunosorbent assay (ELISA) for highly sensitive detection of Hepatitis B virus surface antigen (HBsAg) based on glucose oxidase (GOx)-induced fluorescence quenching of mercaptopropionic acid-modified CdTe quantum dots (MPA-QDs). In this system, hydrogen peroxide (H2O2) sensitive MPA-QDs was used as a signal output, and glucose oxidase (GOx) was used as label which can generate H2O2 via catalytic oxidation of glucose. The proposed method showed dynamic linear detection of HBsAg both in the range of 47pgmL−1 ~ 380pgmL−1 and 0.75ngmL−1 ~ 12.12ngmL−1. The detection limit of the proposed fluorescence ELISA was 1.16pgmL−1, which was approximately 430-fold lower than that of horseradish peroxidase (HRP)-based conventional ELISA. The average recoveries for HBsAg-spiked serum samples ranged from 98.0% to 126.8% with the relative standard derivation below 10%, thus indicating acceptable precision and high reproducibility of the proposed fluorescence ELISA for HBsAg detection. Additionally, the developed method showed no false positive results analyzing 35 real HBsAg-negative serum samples, and exhibited excellent agreement (R2=0.9907) with a commercial time-resolved fluorescence immunoassay (TRFIA) kit for detecting 31 HBsAg-positive serum samples. In summary, the proposed method based on fluorescence quenching of H2O2 sensitive QDs is considerably to be an excellent biodetection platform with ultrahigh sensitivity, good accuracy and excellent reliability.