论文部分内容阅读
用超选择免疫法，诱导ＢＡＬＢ／ｃ小鼠对正常人肝细胞抗原产生选择性低免疫反应后，二步腹腔注射人肝癌细胞悬液，２～４周后用人肝癌细胞作强化免疫动物１次，取出脾细胞与ＳＰ２／０骨髓瘤细胞用ＰＥＧ法融合制备杂交瘤。采用细胞抗原ＥＬＩＳＡ法，同时测定培养上清对肝癌细胞及正常肝细胞的免疫反应，筛出一株能稳定分泌抗人肝癌细胞膜抗原的单克隆抗体（ＨＣＭＰ－１ＭＡｂ）的杂交瘤细胞株。ＨＣＭＰ－１ＭＡｂ是一种ＩｇＧ２ａ亚型的单抗，它与ＨＢｓＡｇ、ＨＢｃＡｇ、ＨＢｅＡｇ等乙型肝炎病毒相关抗原无交叉免疫反应，与ＡＦＰ－亦无阳性反应。免疫萤光法证实ＨＣＭＰ－１ＭＡｂ对ＱＧＹ－７７０３、ＢＥＬ－７４０２及ＳＭＭＣ－７７２１等人肝癌细胞株呈阳性反应。改良ＡＢＣ免疫组织化学证明ＨＣＭＰ－１ＭＡｂ与肝细胞癌组织呈阳性反应。说明ＨＣＭＰ－１ＭＡｂ对肝癌反应的阳性率较高，特导性较好。超选择免疫法，先联合应用免疫抑制药物使动物对正常肝细胞抗原产生低免疫反应性。再以肝癌细胞免疫时，有利于动物免疫系统对肿瘤相关抗原的识别及特异性抗体的产生。从而提高了杂交瘤制备的成功率。我们认为这一方法在单抗制备方面有较高的应用价值。
Superselective immunization was used to induce selective low-immunity responses in BALB/c mice against normal human hepatocyte antigens. Two-step suspensions of human hepatoma cells were injected intraperitoneally, and human hepatocellular carcinoma cells were used as booster immunizations 2 to 4 weeks later. The spleen cells were taken out and SP2/0 myeloma cells were fused with PEG to prepare hybridomas. Cell antigen ELISA was used to simultaneously measure the immune response of the culture supernatant to hepatoma cells and normal hepatocytes, and a hybridoma cell line that can stably secrete a monoclonal antibody (HCMP-1 MAb) against human hepatoma cell membrane antigen was screened out. HCMP-1 MAb is a monoclonal antibody of IgG2a subtype. It has no cross-reaction with hepatitis B virus-related antigens such as HBsAg, HBcAg and HBeAg, and has no positive reaction with AFP-. Immunofluorescence confirmed the positive response of HCMP-1 MAb to human hepatoma cell lines QGY-7703, BEL-7402 and SMMC-7721. Improved ABC immunohistochemistry demonstrated that HCMP-1 MAb is positively reactive with hepatocellular carcinoma. The positive rate of HCMP-1MAb response to hepatocellular carcinoma was higher, and the specific conductivity was better. Superselective immunization methods, combined with the use of immunosuppressive drugs, cause animals to have low immunoreactivity to normal hepatocyte antigens. When hepatocellular carcinoma cells are immunized, the recognition of tumor-associated antigens and the generation of specific antibodies are facilitated by the animal’s immune system. As a result, the success rate of hybridoma preparation is improved. We believe that this method has a high application value in the preparation of monoclonal antibodies.