A comprehensive HBV array for the detection of HBV mutants and genotype

Abstract

Objective: To develop a comprehensive hepatitis B virus (HBV) array providing simultaneous analysis of 8 genotypes, 47 mutations of reverse-transcriptase polymerase gene and 18 mutations of S gene. Method: Oligonucleotides corresponding to various HBV-normal and -mutant sequences were spotted onto pre-treated glass slides. Single-stranded templates of the HBV gene fragment were prepared from serum-DNA of HBV-infected patients by 2-staged PCR and subjected to allele-specific arrayed-primer extension with Cy5-dCTP. Fluorescein-labelled products were scanned at 670 nm. Results: Comparative analysis of 100 unrelated samples using the array and a commercial kit, revealed 44 with additional mutations from the array, these were confirmed by sequencing. Analysis of 381 samples from 45 patients during 1-3 years of anti-viral therapy showed improved sensitivity with detection of drug-resistant mutations months before clinical relapse. The lower detection limit was 28 copies/mL. Conclusion: The array is better than many existing methods as it provides both mutations and genotype data in a single analysis. © 2011 The Canadian Society of Clinical Chemists.