HTLV-I的分离研究

Isolation of Human T-cell Leukemia Virus Type-I

目的建立从HTLV-I感染者中分离HTLV-I的方法。方法采集HTLV-I感染者的肝素抗凝血,分离外周血单核细胞(PBMCs),与健康供者PBMCs或BJAB细胞共培养进行HTLV-I的分离,用P19抗原ELISA试剂盒检测共培养上清,提取培养细胞的基因组DNA,用HTLV-I的Gag、Tax两引物进行PCR扩增,对扩增产物进行琼脂糖凝胶电泳,并进行测序确定病原分离结果。结果从5例经WB确认的HTLV-I感染者中分离到4株HTLV-I毒株,P19抗原显示阳性结果,PCR法也扩增出HTLV-I前病毒DNA,对扩增产物的Gag区进行测序,亦证实该片段为HTLV-I Gag区序列。结论HTLV-I已首次在国内成功分离。

To establish a method to isolate Human F-cell leukemia Virus Type I （HTLV-I） from seropositive individuals. The peripheral blood mononuclear cells（PBMCs） from heparinized venous blood samples of HTLV-I seropositive individuals were collected and cocultured with PBMCs donated by uninfected healthy donors. Supernatant of cocultures was detected by HTLV-I P19 antigen ELISA kit and the proviral DNA of HTLV-I from coculture cells was extracted by using the DNA extractor kit. The amplified products were detected with agarose gel electrophoresis and sequenced to identify the strain of isolation. Of the 5 specimens,4 showed positive for the P19 antigen. The proviral DNAs of specimens were amplified by PCR The am- plified products were sequenced and showed that they were the sequences of HTLV-I. It was evident that HTLV-I virus was successfully isolated firstly in China.