Gelder F B, McDonald J C, Landreneau M D, McMillan R M, Aultman D F
Department of Surgery, Louisiana State University School of Medicine, Shreveport.
Ann Surg. 1991 Jun;213(6):591-7; discussion 597-9. doi: 10.1097/00000658-199106000-00009.
Human lymphocyte antigen (HLA) class I and class II antigens and beta 2 microglobulin (B2M) were identified in peritoneal dialysate (PD) and serum from patients with end-stage renal disease (ESRD) using monoclonal antibodies in an enzyme-linked immunoassay. The HLA class I and class II antigens each exhibited approximate molecular weights of 50,000 to 60,000 daltons by chromatography on Sepharose CL 6B. Class I antigens in serum and PD fluid were associated with B2M. Free B2M (Mr 11,500) also was detected in both sera and PD fluids. Unlike class I antigens, class II antigens were not found to have attached B2M. Class I and class II antigens eluted from 2-diethylaminoethanol ion exchange gradient columns at 0.07 mol/L (molar) phosphate buffer pH 7.2 and migrated with alpha 2-beta 1 mobility in agarose electrophoresis. Class I antigens were purified from ESRD patients' PD fluid by solid-phase immunoaffinity chromatography. Enzyme-linked immunoassay demonstrated that this purified protein was composed of a class I heavy chain and B2M. Class I allospecificity was confirmed by neutralization on known HLA typing antisera in a microcytotoxicity assay. Soluble HLA class I antigen preparations specifically inhibited blast transformation of responder lymphocytes in mixed lymphocyte culture reactions. Inhibition was dose dependent and ranged from 0% to 95%. The presence of soluble HLA antigens in body fluids may play an important part in the immunologic tolerance to self. This study demonstrates a ready source of large quantities of soluble HLA for detailed analysis.
采用酶联免疫分析法，运用单克隆抗体，在终末期肾病（ESRD）患者的腹膜透析液（PD）和血清中鉴定了人类淋巴细胞抗原（HLA）I类和II类抗原以及β2微球蛋白（B2M）。通过在琼脂糖凝胶CL 6B上进行色谱分析，HLA I类和II类抗原各自呈现出约50,000至60,000道尔顿的分子量。血清和腹膜透析液中的I类抗原与B2M相关。在血清和腹膜透析液中也检测到了游离的B2M（分子量11,500）。与I类抗原不同，未发现II类抗原附着有B2M。I类和II类抗原在0.07摩尔/升磷酸盐缓冲液（pH 7.2）中从二乙氨基乙醇离子交换梯度柱上洗脱，并在琼脂糖电泳中以α2-β1迁移率迁移。通过固相免疫亲和色谱法从ESRD患者的腹膜透析液中纯化了I类抗原。酶联免疫分析表明，这种纯化的蛋白质由I类重链和B2M组成。在微量细胞毒性试验中，通过对已知HLA分型抗血清的中和作用证实了I类同种特异性。可溶性HLA I类抗原制剂在混合淋巴细胞培养反应中特异性抑制反应性淋巴细胞的增殖转化。抑制作用呈剂量依赖性，范围为0%至95%。体液中可溶性HLA抗原的存在可能在自身免疫耐受中起重要作用。本研究证明了有大量可溶性HLA可供详细分析的现成来源。
