Dimitrova E, Taskov H, Pashov A
National Center of Infectious and Parasitic Diseases, Sofia, Bulgaria.
Biologicals. 1993 Sep;21(3):215-20. doi: 10.1006/biol.1993.1078.
The present study assesses the influence of the immunophenotyping procedure, the type of monoclonal antibody (MAb) and flow cytometer used, upon the reproducibility of the results of estimation of the normal values for CD3, CD4, and CD8 positive lymphocyte subpopulations. A hundred healthy adults were immunophenotyped by direct immunofluorescence with commercially available (OKT3-FITC, OKT4-PE and OKT8-PE), and locally prepared (HL3-FITC, HL4-PE and HL8-PE)MAb. The samples were analysed on FACStar flow cytometer. The maximal values of the analytical (CD3-0.96, CD4-1.05, CD8-1.02), the individual (CD3-2.33, CD4-2.88, CD8-2.86), and the population (CD3-8.7, CD4-8.8, CD8-7.5) variation prove that the precision of immunophenotyping procedure has no effect on the variation of the values of the CD3+, CD4+ and CD8+ subpopulations. The values of all three markers showed Gaussian type of distribution. Regression analysis proved that the values of these subpopulations determined by MAb from two sources were highly correlated (r > 0.9) but the paired t-test gave statistically significant difference in the values of CD8 (1.23%, P > 0.001). Mean, SD and 95% range of the sizes of the positive subsets determined by OKT3, OKT4 and OKT8 showed that the results were very close to the results from a multicenter study of adult Caucasians. The values we find fall within the cited critical range for these markers; so we can conclude that if good care is taken for the precision of the procedure, MAb source and type of flow cytometer do not influence the values of the assayed subsets.
本研究评估了免疫分型程序、所用单克隆抗体（MAb）的类型和流式细胞仪对CD3、CD4和CD8阳性淋巴细胞亚群正常值估计结果可重复性的影响。采用市售（OKT3-FITC、OKT4-PE和OKT8-PE）和本地制备（HL3-FITC、HL4-PE和HL8-PE）的MAb，通过直接免疫荧光法对100名健康成年人进行免疫分型。样本在FACStar流式细胞仪上进行分析。分析变异（CD3-0.96、CD4-1.05、CD8-1.02）、个体变异（CD3-2.33、CD4-2.88、CD8-2.86）和总体变异（CD3-8.7、CD4-8.8、CD8-7.5）的最大值证明，免疫分型程序的精密度对CD3+、CD4+和CD8+亚群的值的变异没有影响。所有三个标志物的值均呈高斯分布类型。回归分析证明，由两种来源的MAb测定的这些亚群的值高度相关（r>0.9），但配对t检验显示CD8的值存在统计学显著差异（1.23%，P>0.001）。由OKT3、OKT4和OKT8测定的阳性亚群大小的均值、标准差和95%范围表明，结果与成年高加索人的多中心研究结果非常接近。我们发现的值落在这些标志物引用范围内；因此我们可以得出结论，如果对程序的精密度、MAb来源和流式细胞仪类型给予充分注意，则它们不会影响所检测亚群的值。
