Studies on the Development of Viral Detection Markers for the Quality Control of Blood

ABSTRACT

According to the serological screening methods of antigen-antibody reaction such as ELISA, it has been known that the complete detection of viral infections of HBV, HCV, and HIV-1 viruses in the blood and blood related-products is not much reliable. Therefore, nucleic acid amplification testing methods (NAT) adopted to detect the small quantitative viral nucleic acids could support the basis of using and supplying the blood and its related products safely. This research work is basically designed to describe the simultaneous blood screening system by multiplex or duplex tests for detection of HBV, HCV, and HIV-1 viruses in the blood at one time with low price and labor. It is aimed at easy detection by using the conventional agarose gel electrophoresis. Thus, we tried to detect and identify the viral components in the blood sample according to their different size of PCR products. We decided a set of consensus sequences to recognize each viral DNA fragments after running the multiplex PCR in one tube. This was done by nested RT-PCR using two different RNA viral genomic templates followed by multiplex PCR with addition of viral DNA and their primers after purifying the viral genomic nucleic acids. Those specific primers could be used without any interference to amplify each viral genome in the blood samples. The sensitivities with different viral loads were evaluated on the agarose gel electrophoresis. Three different viral agents in the blood samples could be tested by this multiplex (RT)-PCR with three different primers.