流式细胞术计数CD34^(+)细胞的性能验证方法和指标分析

Analytical performance verification protocols and specifications of CD34^(+)cell enumeration by flow cytometry

目的探讨适用于临床实验室进行流式细胞术计数CD34^(+)细胞的性能验证方法和评价标准。方法参考国际指南和我国行业标准,设计流式细胞术计数CD34+细胞(包括相对计数和绝对计数)的性能验证方案,选用全国室间质评(EQA)发放的4个批号质评物,并收集3份干细胞采集物和3份外周血标本,对FACSCantoⅡ配套检测系统的精密度、线性、携带污染率、正确度和准确度实施性能验证,并基于当前技术水平设定评价标准。结果相对、绝对计数的批内CV分别为2.5%~8.9%和3.0%~9.0%,批间CV分别为2.8%~10.5%和3.8%~9.9%;低浓度段(3.6~123.6个/μl)和高浓度段(113.2~1196.3个/μl)线性回归方程的斜率分别为0.9932和0.9652,R2分别为0.9996和0.9939,偏差为-8.67%~0.22%;相对、绝对计数的携带污染率分别为0.07%和0.00%;相对计数≤0.2%或绝对计数≤20个/μl时,正确度绝对偏倚在±0.006%或±0.5个/μl范围内,准确度绝对偏差在±0.02%或±0.9个/μl范围内;相对计数>0.2%或绝对计数>20个/μl时,相对偏倚在±5.65%范围内,相对偏差在±8.19%范围内。各验证结果均在设定的评价标准允许范围内。结论本研究制定的性能验证方案和设定的评价标准,既遵循国内外指南的推荐,又符合我国临床实验室的检测技术水平,具有可操作性,可为临床实验室开展性能验证工作提供参考。

Objective To investigate the analytical performance verification protocols and performance specifications of CD34^(+)cell enumeration by flow cytometry for clinical laboratories.Methods According to international guidelines and National Health Standard of China,we designed the performance verification protocols of CD34+cell enumeration(including percent count and absolute count)by flow cytometry.Four quality assessment materials,three leukapheresis products and three samples of peripheral blood were selected to verify the precision,linearity,carryover,trueness and accuracy of FACSCantoⅡmeasurement system,and the assessment criterion was set according to the detection technologies of clinical laboratories.Results The CVs of intra-run precision of percent count and absolute count were 2.5%to 8.9%and 3.0%to 9.0%;the CVs of inter-run precision were 2.8%to 10.5%and 3.8%to 9.9%,respectively.The slopes of linearity regression equation of low range(3.6/μl to 123.6/μl)and high range(113.2/μl to 1196.3/μl)were 0.9932 and 0.9652,and R^(2) were 0.9996 and 0.9939,and the biases were-8.67%to 0.22%.The carryover of percent and absolute count were 0.07%and 0.00%.When percent count≤0.2%or absolute count≤20/μl,the absolute biases of trueness were in the range of±0.006%or±0.5/μl,and the absolute biases of accuracy were in the range of±0.02%or±0.9/μl;when percent count>0.2%or absolute count>20/μl,the relative biases of trueness were in the range of±5.65%,and the relative biases of accuracy were in the range of±8.19%.The verification results met the assessment criterion set in this study.Conclusions The performance verification protocols and assessment criterion formulated in this study not only conform to the recommendations of domestic and foreign guidelines,but also conform to state of the detection technologies of native clinical laboratories,which can be taken as a reference of performance verification for clinical laboratories.