Development of an amplified luminescent proximity homogeneous assay for quantitative determination of hepatitis B surface antigen in human serum.
BACKGROUND
Hepatitis B virus (HBV) poses a serious risk to human health and the hepatitis B surface antigen (HBsAg) is a popular biomarker in the diagnosis of HBV infection. A quantitative method with a high degree of accuracy, sensitivity and throughput is needed.


METHODS
A novel amplified luminescent proximity homogeneous assay (AlphaLISA) was developed for HBsAg determination. A set of monoclonal antibodies was screened against the main subtypes of HBsAg (adr, ay) to confirm the assay's sensitivity to mutants. Technological processes and reaction conditions were optimized and the assay performance was evaluated.


RESULTS
HBsAg concentrations were determined within a linear range of 0.04 to 100 IU ml(-1). The detection sensitivity was established as 0.01 IU ml(-1). Assay sensitivity to mutant HBsAg was achieved through antibody screening. The results demonstrate that the reproducibility, recovery, and specificity of this assay for HBsAg were better than acceptable. Compared with the commercial light-initiated chemiluminescence assay, the correlation coefficient of the novel assay was established as 0.921.


CONCLUSIONS
The novel AlphaLISA developed in this study has shorter incubation time and easier protocol than the ones of conventional ELISA. It could be used for the clinical determination of HBsAg in human serum. We established a platform for further development of other biomarkers.
Institute of Antibody Engineering, School of Biotechnology, Southern Medical University, Guangzhou 510515, PR China.
Tian-Cai
TC
Liu
He
H
Huang
Zhi-Ning
ZN
Dong
An
A
He
Ming
M
Li
Ying-Song
YS
Wu
Wei-Wen
WW
Xu
D006514:Hepatitis B Surface Antigens / Q000097:blood*; D006801:Humans; D008163:Luminescent Measurements / Q000295:instrumentation / Q000379:methods*
D006514:Hepatitis B Surface Antigens
Amplified luminescent proximity homogeneous assay; Chemiluminescence; Hepatitis B surface antigen; Immunoassays