Hasthorpe S, Stokes K, Rogerson J
Cancer Res. 1985 Oct;45(10):5042-9.
Surface membrane antigen(s) expressed on a mouse mast cell line (FMP1) have also been shown to occur on hemopoietic spleen colony-forming units (CFU-S) and granulocyte/macrophage colony- and erythroid burst-forming cells, using a xeno-antiserum raised against FMP1 cells. This mast cell model has been used to obtain antiserum and large quantities of antigen for the biochemical identification of CFU-S and progenitor cell antigen(s). Immunoprecipitation of FMP1 membrane antigens with the antiserum and subsequent polyacrylamide gel electrophoresis revealed the presence of five membrane proteins with molecular weights of 28,000, 32,000, 36,000, 50,000, and 70,000. Mouse B-lymphoma cell line W279 which reacted with anti-FMP1 serum was found to possess three immunoprecipitable surface proteins with molecular weights of 32,000, 50,000, and 70,000. Attempts have been made to identify the antigen(s) expressed by CFU-S and progenitors which were revealed by immunoprecipitation from the tumor lines. The three lower-molecular-weight proteins (Mr 28,000-36,000) were chosen for initial study. Membrane extracts of FMP1 cells were fractionated on Sephacryl S-200, and selective pools of these antigens were made. Antisera to these pools exhibited complement-dependent cytotoxicity to FMP1 cells, bone marrow CFU-S, granulocyte/macrophage colony-forming cells, and erythroid burst-forming units. These antisera immunoprecipitated Mr 28,000, 32,000, and 36,000 proteins from FMP1 cell membrane extracts but not the Mr 50,000 and 70,000 antigens. The W279 line has only one antigen (Mr 32,000) in the lower-molecular-weight range and is able to absorb anti-CFU-S and anti-progenitor activity, which suggests that it is this antigen which is expressed on hemopoietic cells. In addition, thymocytes react with anti-FMP1 serum, and the Mr 32,000 antigen was immunoprecipitated from thymus cell extracts. Binding studies with concanavalin A, wheat germ agglutinin, and lentil lectin indicated that the Mr 28,000-36,000 proteins were glycoproteins. The apparent molecular weights of these proteins on polyacrylamide gels were not altered by reduction and alkylation and therefore do not contain disulfide-linked subunits.
利用针对小鼠肥大细胞系（FMP1）制备的异种抗血清，已证明在该细胞系上表达的表面膜抗原也存在于造血脾集落形成单位（CFU-S）以及粒细胞/巨噬细胞集落形成细胞和红系爆式集落形成细胞上。这个肥大细胞模型已被用于获取抗血清和大量抗原，以对CFU-S和祖细胞抗原进行生化鉴定。用该抗血清对FMP1膜抗原进行免疫沉淀，随后进行聚丙烯酰胺凝胶电泳，结果显示存在5种膜蛋白，其分子量分别为28,000、32,000、36,000、50,000和70,000。发现与抗FMP1血清发生反应的小鼠B淋巴瘤细胞系W279拥有3种可免疫沉淀的表面蛋白，分子量分别为32,000、50,000和70,000。已尝试鉴定CFU-S和祖细胞表达的、从肿瘤细胞系免疫沉淀中揭示的抗原。选择三种低分子量蛋白（分子量28,000 - 36,000）进行初步研究。FMP1细胞的膜提取物在Sephacryl S - 200上进行分级分离，并制备这些抗原的选择性组分。针对这些组分的抗血清对FMP1细胞、骨髓CFU-S、粒细胞/巨噬细胞集落形成细胞和红系爆式集落形成单位表现出补体依赖性细胞毒性。这些抗血清从FMP1细胞膜提取物中免疫沉淀出分子量为28,000、32,000和36,000的蛋白，但没有沉淀出分子量为50,000和70,000的抗原。W279细胞系在低分子量范围内只有一种抗原（分子量32,000），并且能够吸收抗CFU-S和抗祖细胞活性，这表明造血细胞上表达的就是这种抗原。此外，胸腺细胞与抗FMP1血清发生反应，并且从胸腺细胞提取物中免疫沉淀出了分子量为32,000的抗原。用伴刀豆球蛋白A、麦胚凝集素和扁豆凝集素进行的结合研究表明，分子量28,000 - 36,000的蛋白是糖蛋白。这些蛋白在聚丙烯酰胺凝胶上的表观分子量在还原和烷基化后没有改变，因此不包含二硫键连接的亚基。
