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)。这些研究 有必要确定化学特性、生产地点、 行动机制(S)和意识到TSF需要的机制(S) 由于缺乏适当的安全措施，释放和行动尚未执行 检测技术和稳定、纯化源的TSF的可用性。 本提案中描述的研究将提供新的分析程序和 为进一步研究提供合适的三氟化硫。这样做的目的是 建议是：(1)发展TSF的生物和免疫检测；(2)生产 TSF体外培养，并与不同来源的TSF进行比较； (3)纯化TSF；(4)产生抗TSF抗体；(5)建立TSF的部位 TSF的生产；(6)阐明TSF释放的机理(S) 在体内；(7)建立产生TSF的细胞系；(8)确定变化 TSF处理后巨核细胞倍性变化的研究(9) 体内和体外巨核细胞生成；(10)检测血清中的TSF 有血小板生成障碍的患者。这些目标将是 通过利用各种物理、化学和免疫学 方法：(1)以血小板减少性动物血清为来源 (2)人胚胎肾细胞培养和特异性肿瘤 细胞系将用于体外生产；(3)抗TSF血清将用于 在动物体内饲养，并利用单抗技术；(4) 从各种来源提纯TSF将以高效的方式进行 (5)新的检测技术 器官切除和透析将被用于检测 (6)大鼠巨核细胞生成情况的测定 将研究TSF的有效来源的注射；(7)体外 巨核细胞克隆将从小鼠和人的骨髓中产生 和K562细胞；(8)脾切除、低氧和 ~(89)锶将用于研究TSF与促红细胞生成素的相互作用。 这些研究将提供必要的信息来预测 促血小板生成素的作用、产生部位及作用机制(S) 它在动物身上的需求。此外，还开发了检测程序 TSF、单抗的制备及激素的纯化 将使TSF在血小板患者中的作用研究成为可能 生产紊乱。

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.