Viral Liver Diseases
Copyright ©The Author(s) 2002. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Apr 15, 2002; 8(2): 276-281
Published online Apr 15, 2002. doi: 10.3748/wjg.v8.i2.276
Detection of anti-preS1 antibodies for recovery of hepatitis B patients by immunoassay
Jun Wei, Yu-Qin Wang, Zhi-Meng Lu, Guang-Di Li, Yuan Wang, Zu-Chuan Zhang
Jun Wei, Guang-Di Li, Yuan Wang, Zu-Chuan Zhang, Institute of Biochemsitry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China
Yu-Qin Wang, Zhi-Meng Lu, Department of Clinical virology, Rui-Jin Hospital, Shanghai Second Medical University 200025, Shanghai, China
Author contributions: All authors contributed equally to the work.
Supported by the grants No. KY951-A1-301 and No. KY95T-06-03 from the 9th Five Years Plan Key Research Programs of the Chinese Academy of Sciences.
Correspondence to: Zu-Chuan Zhang, Institute of Biochemsitry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China. zhangzc@sunm.shcnc.ac.cn
Telephone: +86-21-64374430 Fax: +86-21-64338357
Received: June 2, 2001
Revised: October 22, 2001
Accepted: October 29, 2001
Published online: April 15, 2002
Abstract

AIM: To establish a convenient immunoassay method based on recombinant antigen preS1(21-119 aa) to detect anti-preS1 antibodies and evaluate the clinical significance of antibodies in hepatitis B.

METHODS: The expression plasmid pET-28a-preS1 was constructed, and a large quantity of preS1(21-119 aa) fragment of the large HBsAg protein was obtained. The preS1 fragment purified by Ni2+-IDA affinity chromatography was used as coated antigen to establish the indirect ELISA based on streptavidin-biotin system for detection of the anti-preS1 antibodies in sera from HBV-infected patients. For follow-up study, serial sera were collected during the clinical course of 21 HBV-infected patients and anti-preS1 antibodies, preS1 antigen, HBV-DNA and other serological HBV markers were analyzed.

RESULTS: preS1(21-119 aa) fragment was highly expressed from the plasmid pET-28a-preS1 in a soluble form in E. coli (30 mg•L⁻¹), and easily purified to high purity over 90% by one step of Ni2+-IDA-sepharose 6B affinity chromatography. The purity and antigenicity of the purified preS1(21-119 aa) protein was determined by 150 g•L⁻¹ SDS-PAGE, Western blot and a direct ELISA. Recombinant preS1(21-119 aa) protein was successfully applied in the immunoassay which could sensitively detect the anti-preS1 antibodies in serum specimens of acute or chronic hepatitis B patients. Results showed that more than half of 19 acute hepatitis B patients produced anti-preS1 antibodies during recovery of the disease, however, the response was only found in a few of chronic patients. In the clinical follow-up study of 11 patients with anti-preS1 positive serological profile, HBsAg and HBV-DNA clearance occurred in 6 of 10 acute hepatitis B patients in 5-6 mo, and seroconversion of HBeAg and disappearance of HBV-DNA occurred in 1 chronic patients treated with lavumidine, a antiviral agent.

CONCLUSION: The high-purity preS1(21-119 aa) coated antigen was successfully prepared by gene expression and affinity chromatography. Using this antigen, a conveniently detective system of anti-preS1 antibodies in sera was established. Preliminarily clinical trial the occurrence of anti-preS1 antibodies in acute hepatitis B patients suggests the clearance of HBV from serum in a short-term time, and anti-preS1 positive in chronic patients means health improvement or recovery from the disease.

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