FSC/SSC和CD45/SSC分群法流式T淋巴细胞亚群分析的实验研究

Experimental study on the detection of T-lymphocyte subsets in peripheral blood by flow cytometry with FSC/SSC or CD45/SSC gating

目的建立并标准化流式细胞术外周血T淋巴细胞亚群含量测定方法,探讨CD45/SSC和FSC/SSC分群法检测的特点与适用性。方法采用FSC/SSC分群的三色流式细胞术和CD45/SSC分群的四色流式细胞术,对194例健康人外周血T细胞亚群进行测定,并调查两种方法检测结果的精密性及临床应用的成功分群率。结果 FSC/SSC分群法健康人外周血总T细胞、T4细胞、T8细胞、T4/T8比值、DN-T细胞、DP-T细胞的百分含量依次为(69.98±5.79)%、(35.25±5.16)%、(25.08±4.34)%、(6.17±3.08)%、(0.71±0.36)%和(1.45±0.28)%;CD45/SSC分群法结果依次为(71.42±6.02)%、(35.82±5.78)%、(24.98±4.45)%、(5.98±3.11)%、(0.67±0.32)%和(1.47±0.30)%,两种方法检测结果之间差异无统计学意义(F检验,P>0.05)。精密度实验显示,FSC/SSC分群法和CD45/SSC分群法的变异系数均在4%以内,两者之间差异无统计学意义(P>0.05)。临床945份标本分析中,FSC/SSC分群法和CD45/SSC分群法的成功分群率分别为97.78%和99.83%,两者之间差异无统计学意义(P>0.05)。结论 FSC/SSC分群法和CD45/SSC分群法均可用于外周血淋巴细胞亚群含量的检测;FSC/SSC分群法成本相对低廉,上机操作简便,适合大规模临床应用开展,CD45/SSC分群法适合用于特殊标本的复查。

Objective To establish and standardize a method of percentage analysis of T-lymphocyte subsets in peripheral blood by flow cytometry and investigate the character and adaptivity of CD45/SSC and FSC/SSC clustering methods.Methods T-lymphocyte subsets in peripheral blood were detected by three-color flow cytometry of FSC/SSC clustering and four-color flow cytometry of CD45/SSC clustering respectively in 194 health people,then the precision and the clinic successful clustering ratio of the two methods were investigated.Results The percentage of T cell,CD4＋T cell,CD8＋T cell,DN-T cell,DP-T cell and CD4＋T/ CD8＋T,detected by FSC/SSC method,were （69.98±5.79）%,（35.25±5.16）%,（25.08±4.34）%,（6.17±3.08）%,（0.71±0.36）% and （1.45±0.28）%,and those,detected by CD45/SSC method were （71.42±6.02）%,（35.82±5.78）%,（24.98±4.45）%,（5.98±3.11）%,（0.67±0.32）% and （1.47±0.30）%.No significant difference between the results of the two methods（F test,P0.05） was demonstrated.Coefficient of variability in both of the two methods were within 4%,no significant difference was recorded in the precision test（P0.05）.The successful clustering ratio of FSC/SSC and CD45/SSC methods were 97.78% and 99.83% respectively,and there was no statistical difference （P0.05）.Conclusion Both of FSC/SSC and CD45/SSC clustering methods could be used to detect T-lymphocyte subsets in peripheral blood.FSC/SSC method might have the advantage of low cost and simple operation,so it might be fit for large scale clinical application,and CD45/SSC method could be applied to recheck the special samples.