染色体拷贝数变异国家参考品的制备及标定

Preparation and calibration of the national reference for detection of chromosome copy number variations

目的制备染色体拷贝数变异(CNVs)国家参考品,并进行标定。方法收集3份正常人样本,24份染色体非整倍体异常样本,11份微缺失微重复样本,提取基因组DNA,制备成国家参考品。用4家协作单位的染色体拷贝数检测试剂盒对国家参考品性能进行验证,确定准确性,并考察国家参考品在不同条件下(37℃条件下放置4周;放置于37℃条件下4周,期间拿出反复冻融3次)的稳定性及均匀性。结果对43份浓度为15(±1)ng/μL国家参考品的检测结果中,有三家单位均检测出国家参考品标示的结果,有1家单位漏检一例嵌合体样本(1p36 30%)。经不同条件(37℃条件下放置4周;放置于37℃条件下4周,期间拿出反复冻融3次)处理的国家参考品的稳定性结果均一致,均匀性的结果均一致,表明满足参考品的要求。结论成功建立染色体拷贝数变异国家参考品,该参考品可满足对染色体拷贝数检测的要求,经多家实验室进行验证,可用于国内大多数试剂盒的性能评价及临床实验室的质量评价。

Objective To prepare and calibrate the national reference for chromosome copy number variations(CNVs). Methods The national reference was prepared based on DNA extracted from 3 normal cases,24 cases with aneuploidies,and 11 cases with microduplications or microdeletions. Moreover, the accuracy of the national reference was verified by chromosomal copy number variations detection kits from four manufacturers,and the stability and homogeneity under various conditions(storage at 37℃ for 4 weeks;37℃ for 4 weeks and 3 cycles of freeze.thaw at -20℃ and 37℃)were also evaluated. Results All the 43 cases with the concentration of 15(±1)ng/μL in the national reference were accurately detected by the four detection kits except for 1 case(1p36 30%)not detected by one of the kits. Additionally,the national reference treated with different conditions (storage at 37℃ for 4 weeks;37℃ for 4 weeks and 3 cycles of freeze.thaw at - 20℃ and 37℃) exhibited high stability and homogeneity,implying it is qualified to be a reference. Conclusion With the verification of multiple laboratories,the prepared national reference for chromosome copy number variations meets the requirements for the detection of CNVs and is applicable for the evaluation of performances of most domestic detection kits and quality control of clinical laboratories.