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目的研制得到能模拟真实临床标本的人乳头瘤病毒(HPV)高危型16,18核酸检测质控物,并探讨其用于临床实验室室间质量评价的有效性。方法采用分子克隆方法得到含 HPV-16靶基因的重组质粒载体,转染已含 HPV-18的宫颈癌上皮细胞系(HeLa),获得含 HPV-16,18核酸的上皮细胞原液,经适当灭活后甲醇固定。将该细胞原液适当稀释后,组成含阴阳性共12份标本的样本盘,发至全国各开展临床 HPV-16,18检测的实验室,对回报结果进行分析。结果细胞原液经1:10,1:50,1:100,1:500稀释,实时荧光 PCR 检测,HPV-16 Ct 值分别为29.10,31.19,32.15和32.73,HPV-18 Ct 值分别为30.32,32.13,32.22和35.55。质控物在4℃可稳定40 d 以上,盲邮至新疆乌鲁木齐、内蒙古呼和浩特和厦门的样本,返回后检测无明显变化。44个临床实验室对高浓度样本的检测符合率平均为95.1%,对低浓度样本的检测符合率为57.4%。对阴性样本符合率为98.3%。结论获得可模拟临床标本的 HPV-16,18核酸检测质控物,质评结果表明可有效地用于临床实验室室间质评。
OBJECTIVE: To develop a quality control HPV 16 16 nucleic acid test substance that can simulate real clinical specimens and to evaluate the effectiveness of the HPV quality control method in clinical laboratory. Methods The recombinant plasmid vector containing HPV-16 target gene was obtained by molecular cloning and transfected into cervical epithelial cell line (HeLa) containing HPV-18 to obtain the epithelial cell fluid containing HPV-16,18 nucleic acid. Live methanol fixed. The cell stock solution was diluted appropriately to form a sample dish containing a total of 12 positive and negative samples, which were sent to laboratories that conducted clinical tests of HPV-16 and HPV 18 in the whole country to analyze the return results. Results The results of real-time PCR showed that the Ct values of HPV-16 were 29.10, 31.19, 32.15 and 32.73, respectively. The HPV-18 Ct values were 30.32, 32.13, 32.22 and 35.55. The quality control material can be stable for more than 40 days at 4 ℃. After blind mail to Urumqi, Xinjiang, Hohhot, Inner Mongolia and Xiamen samples, there is no obvious change after the test. The average coincidence rate of 44 clinical laboratories for high-concentration samples was 95.1%, while that for low-concentration samples was 57.4%. The coincidence rate of negative samples was 98.3%. Conclusion The HPV-16, 18 nucleic acid quality control test samples that can be used as clinical samples were obtained. The results of qualitative assessment indicated that HPV-16,18 could be used to evaluate the clinical laboratory quality effectively.