Purification of Common Lung Cancer Antigen recognized with Monoclonal Antibody TFS and Production of Monoclonal Antibodies of Second Generation by its antigen.

制备了抗小细胞肺癌单克隆抗体（MAb）TFS-2，并与所有组织类型的肺癌发生反应。最近，我们尝试用^<99m>Tc标记TFS-2对人肺癌进行放射免疫检测，然后在骨、肺、肝等各种组织中获得原发性和转移性肺癌的清晰成像。(1)肺癌常见抗原的纯化。以转移到肝脏的小细胞肺癌肿块为纯化源。样品用0.5% NP-40的PBS溶解。样品经过两次亲和层析（三烷基活化的Sepharose 4B偶联MAb TFS-2）。用乙醇-沉淀法去除NP-40。SDS-PAGE显示2条蛋白带，分别为39kDa和33kDa。Western blotting上33kda条带与MAb TFS-2反应。33kDa的蛋白直接转移到膜上分析，并测定了20个氨基酸残基。(2)肺癌常见抗原基因编码分析。该MRNA从小细胞肺癌NCI-1169细胞系中获得，构建质粒表达CDNA文库。将质粒转染到Cos-1细胞中，用^<125>I标记的MAb TFS-2筛选细胞表面表达抗原的克隆。分离到了一个编码该蛋白的EDNA克隆，该蛋白与单抗TFS-2反应。(3)第二代单克隆抗体的产生目前已获得几种能与纯化抗原反应的杂交瘤细胞克隆。

Monoclonal antibody(MAb)TFS-2 was produced against Small Cell Lung Cancer and reacted with all tissue types of lung cancer. Recently we tried the radioimmunodetection of human lung cancer with ^<99m>Tc labeled TFS-2 followed by getting clear imaging of primary and metastatic lung cancers in the various tissues such as bone, lung, and liver.(1)Purification of Common Lung Cancer Antigen.Masses of metastatic small cell lung cancer to liver was used as source for purification. Samples were solubilized by PBS with 0.5% of NP-40. Samples were twice passed through Affinity Chromatography(tresyl-activated Sepharose 4B couppled with MAb TFS-2). NP-40 was removed by ethanol pi-ecipitatiori. SDS-PAGE showed 2 bands of protein, 39kDa and 33kDa respectedly. Band of 33 kDa reacted with MAb TFS-2 on the Western blotting. Protein of 33kDa was analyzed directly transferring to membrane and 20 amino acid residue was determined.(2)Analysis of Gene coding Common Lung Cancer Antigen.The MRNA was obtained from small cell lutig cancer cell line NCI-1169, and the expression CDNA library in plasmid was constructed from the MRNA. The plasmids were transfected to Cos-1 cells and clones expressing antigen on the cell surface were screened by ^<125>I labeled MAb TFS-2. A EDNA clone coding the protein reacted with MAb TFS-2 was isolated.(3)Production of Monoclonal antibodies of the second generation Several clones of hybridoma cell that reacted with purified antigen presently were obtained.