Maher Kevin J, Klimas Nancy G, Hurwitz Barry, Schiff Richard, Fletcher Mary Ann
Department of Medicine, University of Miami School of Medicine, Miami, Florida 33136, USA.
Clin Diagn Lab Immunol. 2002 Nov;9(6):1248-52. doi: 10.1128/cdli.9.6.1248-1252.2002.
We present methodologic details and operating characteristics of a procedure with whole blood for the quantitative assessment of intracellular perforin within distinct lymphocyte subsets. Using this method, we analyzed 20 healthy controls and 2 individuals with an inherited deficiency of perforin. The mean +/- standard deviation perforin contents of natural killer (NK) cells and cytotoxic T cells of healthy controls were 3561 +/- 1157 and 500 +/- 779 relative number of molecules (rMol) of antiperforin antibody bound per cell, respectively. The NK cell perforin contents of individuals with heterozygous and homozygous perforin deficiency (familial hemophagocytic lymphohistiocytosis) were 2260 and 212 rMol of antiperforin antibodies per NK cell. While the homozygous deficiency was found to be associated with negligible antiperforin binding, the heterozygous condition was associated with a level of perforin binding that was below the 15th percentile for healthy individuals. Because 83% of this subject's NK cells were shown to bind to antiperforin antibodies by conventional flow cytometry (relative to the normal range of 81% +/- 25%), quantitative cytometry may be more sensitive than conventional cytometric methods in identifying cytolytic defects.
我们介绍了一种使用全血对不同淋巴细胞亚群内细胞穿孔素进行定量评估的方法的详细方法学和操作特性。使用该方法，我们分析了20名健康对照者和2名遗传性穿孔素缺乏症患者。健康对照者自然杀伤（NK）细胞和细胞毒性T细胞的平均±标准差穿孔素含量分别为每细胞3561±1157和500±779个抗穿孔素抗体结合分子的相对数量（rMol）。杂合子和纯合子穿孔素缺乏症（家族性噬血细胞性淋巴组织细胞增生症）患者的NK细胞穿孔素含量分别为每NK细胞2260和212 rMol抗穿孔素抗体。虽然发现纯合子缺乏与可忽略不计的抗穿孔素结合有关，但杂合子情况与穿孔素结合水平低于健康个体的第15百分位数有关。由于通过传统流式细胞术显示该受试者83%的NK细胞与抗穿孔素抗体结合（相对于81%±25%的正常范围），定量细胞术在识别溶细胞缺陷方面可能比传统细胞术方法更敏感。
