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Application of the NucliSens Basic Kit for detection of Hepatitis C Virus

Application of the NucliSens Basic Kit for detection of Hepatitis C Virus

Abstracts of the General Meeting of the American Society for Microbiology 101: 179

Background and Aim: The Hepatitis C Virus (HCV) is a major cause of acute and chronic liver disease. The aim of this study was to develop an assay for HCV detection based on nucleic acid sequence based amplification (NASBA) using the NucliSens(R) Basic Kit (Organon Teknika) and to evaluate this technique with clinical specimens. Methods: The assay utilized kit-based reagents for nucleic acid release, "Boom" silica-based nucleic acid extraction, isothermal NASBA amplification and ECL (electrochemiluminescence) detection. These reagents were used in combination with primers and probes designed within the 5' non-coding region of the HCV genome. Plasma samples (n=55) submitted for HCV diagnosis or HCV viral load were analyzed with the Basic Kit. Basic Kit results were evaluated in comparison with a well established qualitative "in-house" reverse transcription-nested PCR (RT-nPCR) and with commercial HCV quantitative assays. The sensitivity of the Basic Kit assay was estimated by testing serial dilutions of HCV positive plasma samples of known viral load. Results: The results obtained for HCV detection in plasma specimens were: 18/46 Basic Kit positive and 17/46 RT-nPCR positive. The remaining nine samples, all with detectable HCV viral load, were also positive by Basic Kit. The Basic Kit assay was capable of detecting approximately 100 copies of HCV RNA input. Conclusions: The performance of the Basic Kit assay for detection of HCV in clinical specimens was comparable to the "in-house" RT-nPCR method. Positive Basic Kit results also correlated with detectable HCV viral load in clinical samples. The sensitivity of the Basic Kit-based HCV assay was estimated to be 100 copies of input. The NucliSens Basic Kit could become a useful diagnostic tool for NASBA-based HCV detection since it offers good sensitivity and short turnaround time.

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