Research On The Visual Gene Microarray For Detection Of Hepatitis A Virus Using Gold-DNA Probes

Hepatitis A virus (HAV), a major cause of acute hepatitis infections in humans in many countries, is a positive strand RNA virus belonging to the picornavirus family. HAV infection occurs primarily by the fecal-oral route, by person-to-person contact or through the ingestion of contaminated food or water. Thus, it is essential to develop special and efficacious methods for the detection of HAV in clinical or biological samples. With the features of high- sensitivity, high- specificity and high-throughput, gene microarray technology has a promising prospect in the rapid diagnosis of clinical disease, and environmental monitoring. Currently a variety of gene chip technologies are commonly applied in medical field, however, they also have demerits in low sensitivity, too complicated technique in detection, or requisition for special conditions and equipments.we (?) have combined organically gene technology with nanotechnology to fully take the advantage of 106 fold of signal amplification by the interaction between nano-gold and silver enhancement and have developed a gene microarray technology for detection of HAVusing gold -DNA Probes. In recent years, the application of gold nanoparticles as oligonucleotide labels instead of fluorescence dyes and enzyme-conjugation in DNA detection assays was very common. Many experiments revealed that gold- labeled probes were compatible with gene microarray or gene-chip and would be used as an alternative marker because of their high stability, high labelling density and superior sensitivity.The principle of HAVgene array: Amino- modified oligodeoxynucleotides at the 5' position were arrayed on an activated glass surface to function as capture probes. Sandwich hybridization occurred among capture probes, the HAV amplicon, and gold nanoparticle-supported oligonucleotide probes. After a silver enhancement step, signals were detected by a standard flatbed scanner or just by naked eyes.Our lab has already reported simultaneous diagnosis of HBV and HCV by using gold nanoparticle probe and silver enhancement. Based on these works, we established HAVdetection method with gold—DNA probe coupling silver enhancement on an array. The concentration of capture probe and gold—DNA probe, silver enhancement time and blocking strategy etc were optimized. The sensitivity of the HAV array was also being measured, the lOOfM HAV amplicon was markedly detectable in a 12min silver enhancement.Therefore, the experiments mentioned above proved that the merits of this gene microarray technology were high sensitivity, excellent specificity, low-cost and easy operation. Meanwhile it could meet the demand of design and hence had a promising prospect in clinical diagnosis <>...