肾移植流式细胞术交叉配型中抗体结合与细胞毒性的同步检测

Simultaneous detection of antibody binding and cytotoxicity in flow cytometry crossmatch for renal transplantation.

作者信息

Won Dong Il, Jeong Hee Du, Kim Yong Lim, Suh Jang Soo

机构信息

Department of Laboratory Medicine, Kyungpook National University Hospital, Daegu, Korea.

出版信息

Cytometry B Clin Cytom. 2006 Mar;70(2):82-90. doi: 10.1002/cyto.b.20089.

DOI:10.1002/cyto.b.20089

PMID:16470535

Abstract

BACKGROUND

The anti-HLA antibody can be detected using either a complement-dependent lymphocytotoxicity (CDC) assay or a flow cytometry crossmatch (FCXM) in renal transplantation. Discordant results are often obtained because the two methods detect different reaction phases between the donor lymphocytes and the recipient sera. This study was intended to confirm that antibody binding and cytotoxicity to the lymphocytes can be detected simultaneously in a single FCXM assay, cytotoxic FCXM.

METHODS

In the cytotoxic FCXM, the antibody binding to the lymphocytes was measured using anti-IgG-FITC, and the cytotoxicity using 7-aminoactinomycin D (7-AAD) after adding complement. For an evaluation of two test parameters, the cytotoxicity test moiety (dead-cell percentage) was compared with the anti-human globulin (AHG)-CDC, and the antibody-binding test moiety (sample/control fluorescence ratio) with the conventional FCXM in 77 positive and 30 negative crossmatches.

RESULTS

In the cytotoxic FCXM, both antibody binding and cytotoxicity could be assessed in a single anti-IgG-FITC/7-AAD plot. Regarding the correlation between the presence of HLA antibodies and the test result, the cytotoxicity parameter (r = 0.55) appeared to be more suitable than that of the AHG-CDC (r = 0.50) but the antibody-binding parameter (r = 0.83) was worse than that of the conventional FCXM (r = 0.93). The sensitivity of both parameters of the cytotoxic FCXM was not significantly different from each conventional counterpart (P = 0.33 and P = 0.22, respectively).

CONCLUSIONS

The simultaneous detection of Ab binding and cytotoxicity was possible by the cytotoxic FCXM with the test efficiencies similar to the conventional counterparts. If this new assay is improved through the further studies to optimize the critical assay variables, this may be used as an alternative to the conventional assays to acquire more information on the characteristics of the recipient's HLA alloantibodies.

摘要

背景

在肾移植中，可使用补体依赖淋巴细胞毒性（CDC）试验或流式细胞术交叉配型（FCXM）检测抗人白细胞抗原（HLA）抗体。由于这两种方法检测的是供体淋巴细胞与受体血清之间不同的反应阶段，所以常常会得到不一致的结果。本研究旨在证实，在单一的细胞毒性流式细胞术交叉配型（细胞毒性FCXM）试验中可同时检测抗体与淋巴细胞的结合及细胞毒性。

方法

在细胞毒性FCXM中，使用抗IgG-异硫氰酸荧光素（FITC）检测抗体与淋巴细胞的结合情况，加入补体后使用7-氨基放线菌素D（7-AAD）检测细胞毒性。为评估两个检测参数，在77例阳性和30例阴性交叉配型中，将细胞毒性检测部分（死细胞百分比）与抗人球蛋白（AHG）-CDC进行比较，将抗体结合检测部分（样本/对照荧光比率）与传统FCXM进行比较。

结果

在细胞毒性FCXM中，可在单一的抗IgG-FITC/7-AAD图中评估抗体结合和细胞毒性。关于HLA抗体的存在与检测结果之间的相关性，细胞毒性参数（r = 0.55）似乎比AHG-CDC的参数（r = 0.50）更合适，但抗体结合参数（r = 0.83）比传统FCXM的参数（r = 0.93）要差。细胞毒性FCXM两个参数的敏感性与各自传统方法相比无显著差异（分别为P = 0.33和P = 0.22）。