甲型肝炎病毒的纯化及其单抗的制备

应用不连续蔗糖／甘油密度梯度超速离心的方法，从培养细胞中纯化甲肝病毒（Ｈｅ－ｐａｔｉｔｉｓＡＶｉｒｕｓ，ＨＡＶ）。纯化的ＨＡＶ抗原经ＳＤＳ－ＰＡＧＥ电泳分析，获得三条带：ＶＰ１（３４０００），ＶＰ２（３００００）和ＶＰ３（２６０００）。用纯化的ＨＡＶ抗原免疫Ｂａｌｂ／ｃ小鼠，将已免疫小鼠的脾细胞与ＳＰ２／０细胞融合，获得６株分泌抗ＨＡＶ单克隆抗体的杂交瘤。它们的培养上清和腹水的抗体滴度分别是５０～１０００和１０００～４０００。Ｉｇ亚类５株为ＩｇＧ１，１株为ＩｇＧ２ａ。采用间接的ＥＬＩＳＡ法分析ＨＡＶ抗原的抗原位点，结果表明ＨＡＶ抗原至少存在４个抗原位点，两个存在于ＶＰ１蛋白带，另两个分别存在于ＶＰ２和ＶＰ３蛋白带     

Analysis of human class II antigens by cloned cytolytic T cell reagents: a study using HLA loss mutant lymphoblastoid cell lines and monoclonal antibodies detecting the HLA-DP product(s)

We have utilized cloned T cell reagents and ionizing radiation-induced mutants of an HLA heterozygous lymphoblastoid cell line (LCL) to investigate the determinants detected by the cell-mediated lympholysis (CML) assay. Cells of an LCL clone, 721.501, an HLA haplotype loss mutant expressing the HLA-A2-Cw1-Bw51-DR1-Dw1-DQw1-DPw2-GLO haplotype were used as sensitizing cells for responder cells in vitro. "Cloned" reagents were generated by single-cell deposition of cells of a bulk reagent primed against 721.501 cells. Those clones were screened for cytolytic activity against HLA loss mutant targets (derived from LCL 721) of four different categories; HLA-A2 loss only, A2-Cw1-Bw51 loss, A2-Cw1-Bw51-DR1-DQw1 loss, and the entire HLA haplotype loss. Of 196 clones tested, 36 were cytolytic, including three anti-A2, five anti-Bw51/Cw1, 12 anti-DR1/DQw1, 13 anti-DP region associated with DPw2, and three of undetermined specificity, based on cytolytic patterns against the HLA loss mutant targets. Of 25 anti-HLA class II lytic clones, 23 (92%) fitted the characteristics of helper cell-independent cytolytic T cells (HITc), whereas only two of eight (25%) anti-class I clones were HITc. The 13 anti-DP region clones were divided into three subgroups defined by blocking by anti-FA and not Tü39 monoclonal antibodies (MoAb), by Tü39 and not anti-FA, and by both MoAbs.