转移性体内迟发型超敏反应监测法的建立及临床应用

目的 建立体内转移性迟发型超敏反应(Trans-vivo DTH)检测方法,用于评价临床移植受者的免疫状态.方法 通过Ficoll密度梯度离心法获取供者和受者(各13例)外周血单个核细胞(PBMC),用超声粉碎供者PBMC,制备供者亚细胞抗原.将受者的PBMC、回忆性抗原EBV抗原和供者抗原按以下各组分别注射入CB-17 SCID小鼠的足垫:①阴性对照组:7~9×106个受者PBMC;②供者抗原反应组:7~9×106个受者PBMC+10 μg供者抗原;③阳性对照组:7~9×106个受者PBMC+10 μg EBV抗原;④连锁免疫抑制组:7~9×106个受者PBMC+10 μg EBV抗原+10 μg供者抗原.分别于注射前及注射24 h后双盲测量小鼠的足垫厚度,计算两次测量厚度的差值.实验的有效性依据为阳性对照组足垫净厚度改变≥63.5×10-3 mm.结果 13例亲属活体肾移植受者接受Trans-vivo DTH检测显示,致敏型8例(62%),调节型3例(23%),非调节型2例(15%).结论 Trans-vivo DTH检测法可用于监测移植后受者对供者抗原的反应性.

Establishment of Trans-vivo delayed type hypersensitivity assay and its clinical application

Objective To establish trans-vivo delayed type hypersensitivity (trans-vivo DTH) assay for evaluation of immune response status in renal transplant recipients. Methods Peripheral blood mononuclear cells (PBMC) were obtained from donors (n=13) and recipients (n=13) by Ficoll density gradient centrifugation. Sub-cell antigens were prepared by ultrasonication of PBMC from donors. PBMC from recipients, recall antigen EBV antigens and donor antigens were injected into the foot pads of CB-17 SCID mice accordingly. For negative control group, 7-9×106 PBMC from recipients were injected. For donor antigen reactive group, 7-9×106 PBMC from recipients and 10 μg donor antigens were injected. For positive control group, 7-9 ×106 PBMC from recipients and 10 μg EBV antigens were injected. For linked immune suppression group, 7-9×106 PBMC from recipients, 10 μg EBV antigens and 10 μg donor antigens were injected. The thickness of foot pads of mice was measured before injection and 24 h after injection, and the change of thickness of foot pads was calculated. The net change of thickness ≥ 63.5×10-3 mm in positive control group was considered to be valid. Results Trans-vivo DTH assay in monitoring of 13 living related renal transplant recipients revealed that 8 patients (62%) were sensitized pattern, 3 patients (23%) were regulator pattern, and the other 2 patients (15%) were non-regulator pattern. Conclusion Response to donor antigens in living related renal transplant recipients can be monitored by trans-vivo DTH assay.