Crispr-Cas12a法快速检测脑胶质瘤异柠檬酸脱氢酶1基因突变的效能研究

Efficacy of Crispr⁃Cas12a in rapid detection of isocitrate dehydrogenase 1 gene mutation in glioma

目的运用Crispr-Cas12a法快速检测冰冻脑胶质瘤组织样本中异柠檬酸脱氢酶1(IDH1)基因R132H单核苷酸多态性,初步评价该方法的检测效能及其与直接测序方法的一致性。方法选择30例脑胶质瘤冰冻组织样本,分别使用Crispr-Cas12a法、免疫组织化学法(IHC)以及直接测序法对样本IDH1-R132H位点突变情况进行检测,计算Crispr-Cas12a法的灵敏度、特异性等效能指标,并利用配对χ^(2)检验(McNemar检验)及Kappa一致性检验分析Crispr-Cas12a法、IHC法与金标准(直接测序方法)的一致性。结果利用Crispr-Cas12a法初步实现了60 min内对冰冻脑胶质瘤组织样本IDH1-R132H位点突变的快速检测。以直接测序为金标准,Crispr-Cas12a法的灵敏度为87.7%,特异性为97.0%,一致性为91.1%,Kappa一致性检验显示两种方法一致性较好(k=0.816)。结论Crispr-Cas12a法能快速、准确地对冰冻脑胶质瘤组织样本IDH1-R132H突变进行检测,且稳定性较好,有望成为一种检测IDH1突变的术中检测方法。

Objective The Crispr⁃Cas12a method was used to rapidly detect the R132H single nucleotide polymorphism of the isocitrate dehydrogenase 1(IDH1)gene in frozen glioma tissue samples,and the detection efficiency of this method and its consistency with the direct sequencing method were preliminarily evaluated.Method Thirty cases of glioma frozen tissue samples were selected,and the IDH1⁃R132H mutation was detected by Crispr⁃Cas12a method,immunohistochemistry(IHC)and direct sequencing method respectively.The sensitivity and specificity of Crispr⁃Cas12a method were calculated,and the consistency of Crispr⁃Cas12a method,IHC method and the gold standard(direct sequencing method)was analyzed by pairedχ^(2)test(McNemar test)and Kappa consistency test.Result The rapid detection of IDH1⁃R132H mutation in frozen glioma tissue samples within 60 min was preliminarily realized by Crispr⁃Cas12a method.With direct sequencing as the gold standard,the sensitivity of the Crispr⁃Cas12a method was 87.7%,the specificity was 97.0%,and the consistency was 91.1%.The Kappa consistency test showed that the two methods had good consistency(k=0.816).Conclusion Crispr⁃Cas12a method can quickly and accurately detect IDH1⁃R132H mutation in frozen glioma tissue samples,and has good stability,which is expected to become an intraoperative detection method for IDH1 mutation.