Fluorescent reverse transcription-PCR assay for determination of hepatitis C virus genotypes
目的应用荧光RT-PCR方法对慢性丙型肝炎（CHC）患者血清进行HCV基因型分析。方法采用荧光RT-PCR方法与测序方法比较,对118例CHC阳性患者血清进行HCV RNA分型,比较两种方法分型结果的一致性;为了考察该方法的特异性,对98例慢性乙型肝炎（CHB）患者血清进行HCV RNA分型检测。结果在118例HCV RNA阳性血清标本中,用荧光RT-PCR方法检测有HCV基因1型79例（66.9%）,2型29例（24.6%）,1/2混合型8例（6.8%）;另有2例（1.7%）HCV RNA定量为1×103IU/ml弱阳性标本未分出型。测序法分出1型81例（68.4%）,2型29例（24.6%）,1/2混合型6例（5.1%）,与荧光RT-PCR方法一致2例（1.7%）HCV RNA定量结果为1×103IU/ml的阳性标本测序法未分出型。未发现其他基因型的病例。98例CHB患者血清检测结果均为阴性。结论荧光HCV基因分型的方法具有良好的敏感性、特异性、可重复性且省时省力,可选择该方法为临床诊断服务。Abstract:
Objective To analyze the genotype of blood borne hepatitis C virus (HCV) using fluorescent reverse transcription-PCR (RT-PCR) method.Methods Fluorescent reverse transcription polymerase chain reaction (RT-PCR) detection of HCV RNA was used for the genotype determination of 118 serum samples from HCV patients and compared with sequncing assay of 98 clinical samples from HBV patients checked as negative control.Results Fluorescent RT-PCR result from 118 samples of HCV patients showed 79 (66.9%) were genotype 1, 29 (24.6%) were genotype 2, 8 (6.8%) were genotype 1 and genotype 2 mixed, While 2 (1.7%) samples were non-reactive because HCV RNA was less than 1×103IU/ml.Sequence assay showed, 81 (68.4%) were genotype 1, 29 (24.6%) were genotype 2, 6 (5.1%) were genotype 1 and genotype 2 mixed, 2 (1.7%) were non-reactive also.In control group the samples from HBV patients were negative for HCV genotype detection.Conclusion The fluorescent RT-PCR was sensitive, specific, repeatable and convenient for HCV genotyping and could be used for clinical genotyping.
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