carbocyanines and Hepatitis-B

An aptamer-based competitive binding assay for one-step (i.e. no requirement of pre-treatment) quantitation of target molecules of interest has been developed. This method has been successfully employed for the fast and sensitive detection of the surface antigen of the hepatitis B virus (HBsAg). The key features of our method include its low intrinsic background noise, low costs, high resolution, and high sensitivity, enabling the detection of as low as 1.25 mIU mL(-1), approximately 40-fold better than that of the most widely used Abbott Architect assay for HBsAg detection, without the tedious extraction and/or washing procedures. Moreover, this assay has better recovery and accuracy than that of conventional competitive binding assay or others for HBsAg quantitation.

Topics: Aptamers, Nucleotide; Binding, Competitive; Carbocyanines; Fluorescence Resonance Energy Transfer; Hepatitis B; Hepatitis B Surface Antigens; Hepatitis B virus; Humans; Limit of Detection

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.. 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 670nm.. 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.. The array is better than many existing methods as it provides both mutations and genotype data in a single analysis.

Topics: Carbocyanines; Deoxycytosine Nucleotides; Drug Resistance, Viral; Follow-Up Studies; Genotype; Hepatitis B; Hepatitis B virus; Humans; Limit of Detection; Mutation; Oligonucleotide Array Sequence Analysis; Polymerase Chain Reaction; RNA-Directed DNA Polymerase; Sensitivity and Specificity; Viral Envelope Proteins