THROMBOPOIETIN: IMMUNOASSAY AND CHARACTERIZATION

现在似乎很清楚，血小板的产生是由一种 血小板生成刺激因子（TSF或血小板生成素）。 研究 必须确定化学特性，生产地点， 行动机制和感觉到需要TSF的机制 由于缺乏适当的 分析技术和TSF的稳定、纯化来源的可用性。 本提案中描述的研究将提供新的分析程序和 提供合适的TSF以供进一步研究。 这一目标 建议：（1）开发TSF的生物和免疫测定法;（2）生产 并与不同来源的TSF进行比较; (3)纯化TSF;（4）产生TSF抗体;（5）建立 TSF的产生;（6）阐明导致TSF释放的机制 在体内;（7）建立产生TSF细胞系;（8）测定变化 TSF处理后巨核细胞的倍性;（9）研究 体内和体外巨核细胞生成;（10）检测血清中的TSF 血小板生成障碍的患者。 这些目标将 通过利用各种物理、化学和免疫学方法， 方法：（1）以血吸虫病动物血清为来源 （2）人胚肾细胞培养和特异性肿瘤细胞培养 细胞系将用于体外生产;（3）抗TSF血清将 在动物中饲养和使用单克隆抗体技术;（4） 从各种来源的TSF的纯化将通过高性能 液相色谱和免疫学技术;（5）新的检测技术 和器官切除后透析将用于测定 产生TSF的位点;（6）巨核细胞生成的测量 将研究TSF的有效来源的注射;（7）体外 巨核细胞集落将从鼠和人骨髓中产生 和K562细胞;和（8）脾切除术，缺氧， 89 Sr将用于研究TSF和促红细胞生成素的相互作用。 这些研究将提供必要的信息，以预测模式， 促血小板生成素的作用、产生部位和感觉机制 动物的需要。 此外，还开发了用于 TSF，单克隆抗体的生产和激素的纯化 将使研究TSF对血小板减少患者的作用成为可能。 生产障碍。

It now seems clear that platelet production is controlled by a thrombocytopoiesis-stimulating factor (TSF or thrombopoietin). The studies necessary to determine the chemical characteristics, site of production, mechanism(s) of action, and mechanism(s) that sense the need for TSF release and action have not been performed because of the lack of suitable assay techniques and availability of a stable, purified source of TSF. Research described in this proposal will provide new assay procedures and a suitable supply of TSF for further studies. The objectives of this proposal are to: (1) develop bio- and immunoassays for TSF; (2) produce TSF in vitro and make comparisons to TSF produced from different sources; (3) purify TSF; (4) produce antibodies to TSF; (5) establish the site of production of TSF; (6) clarify the mechanism(s) that cause release of TSF in vivo; (7) establish cell lines that produce TSF; (8) determine changes in ploidy of megakaryocytes after TSF treatment; (9) investigate megakaryocytopoiesis both in vivo and in vitro; and (10) test TSF in sera of patients with platelet production disorders. These objectives will be pursued by utilizing a variety of physical, chemical, and immunological methods: (1) sera from thrombocytopenic animals will be used as a source of TSF; (2) both human embryonic kidney cell cultures and specific tumor cell lines will be utilized for in vitro production; (3) anti-TSF sera will be raised in animals and by use of monoclonal antibody technology; (4) purification of TSF from various sources will be made by high performance liquid chromatography and immunologic techniques; (5) new assay techniques and organ removal followed by dialysis will be used for determination of site(s) of production of TSF; (6) measurement of megakaryocytopoiesis after injection of potent sources of TSF will be investigated; (7) in vitro megakaryocytic colonies will be produced from murine and human bone marrow and K562 cells after stimulation of TSF; and (8) splenectomy, hypoxia, and 89Sr will be used for the study of TSF and erythropoietin interactions. These studies will provide the necessary information to predict the mode of action of thrombopoietin, site of production, and mechanism(s) that sense its need in animals. In addition, the development of assay procedures for TSF, production of monoclonal antibodies and purification of the hormone will make possible studies of TSF action in patients with platelet production disorders.