Monoclonal antibody 9C4 recognizes epithelial cellular adhesion molecule, a cell surface antigen expressed in early steps of erythropoiesis
Monoclonal antibody 9C4 recognizes epithelial cellular adhesion molecule, a cell surface antigen expressed in early steps of erythropoiesis
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DOI：
10.1016/s0301-472x(02)00798-1
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发表时间：
2002-06-01
影响因子：
2.6
通讯作者：
Bühring, HJ
中科院分区：
文献类型：
--
作者：
Lammers, R;Giesert, C;Bühring, HJ
目标.单克隆抗体（mAb）9 C4检测未成熟红系祖细胞和上皮肿瘤细胞系上表达的表面抗原。本研究的目的是鉴定所识别的表面抗原，并分析该分子在红细胞生成的早期步骤中的潜在作用。垂体来源的逆转录病毒cDNA文库用于产生病毒并感染NIH-3 T3成纤维细胞。转染细胞用nAb 9 C4染色;通过FACS分选阳性细胞;并建立结合mAb 9 C4的克隆细胞系。通过聚合酶链反应扩增编码9 C4结合蛋白的cDNA并克隆。用流式细胞仪分析mAb 9 C4与人骨髓细胞的反应性。分离的cDNA序列分析发现与上皮细胞粘附分子（Ep-CAM）的100%的同一性。双色流式细胞术分析显示，几乎100%的Ep-CAM（+）BM细胞共表达CD 105、E-cadherin和高水平的CD 71。Ep-CAM（+）BM细胞的部分也是CD 34（+），但缺乏血型糖蛋白A表达，表明Ep-CAM（+）BM细胞的部分也是CD 34（+），但缺乏血型糖蛋白A表达。细胞代表未成熟的红系细胞。骨髓单个核细胞的逆转录聚合酶链反应分析显示，9 C4（+）成红细胞群体而不是9 C4（-）部分表达Ep-CAM mRNA。体外诱导分化为红系的外周血CD 34（+）细胞在培养的第3至7天显示出强的Ep-CAM表达。加入Ep-CAM特异性单克隆抗体9 C4或KS 1/4可使Ep-CAM蛋白表达上调2 - 3倍。mAb 9 C4识别Ep-CAM，一种在红细胞生成早期阶段表达的分子。(C)2002年国际实验血液学学会。出版社：Elsevier Science Inc.
Objectives. Monoclonal antibody (mAb) 9C4 detects a surface antigen expressed on immature erythroid progenitor cells and epithelial tumor cell lines. The aim of this study was to identify the recognized surface antigen and to analyze a potential role of this molecule in early steps of erythropoiesis.Materials and Methods. A pituitary-derived retroviral cDNA library was used to generate viruses and infect NIH-3T3 fibroblasts. The transfected cells were stained with nAb 9C4; positive cells were sorted by FACS; and a clonal cell line binding mAb 9C4 was established. cDNA encoding the 9C4-binding protein was amplified by polymerase chain reaction and cloned. Reactivity of mAb 9C4 with human bone marrow (BM) cells was analyzed by flow cytometry.Results. Sequence analysis of the isolated cDNA uncovered a 100% identity with the epithelial cellular adhesion molecule (Ep-CAM). Two-color flow cytometric analysis revealed that almost 100% of Ep-CAM(+) BM cells coexpressed CD105, E-cadherin, and high levels of CD71. Fractions of Ep-CAM(+) BM cells also were CD34(+) but lacked glycophorin A expression, suggesting that Ep-CAM. cells represent immature erythroid cells. Reverse transcriptase polymerase chain reaction analysis of BM mononuclear cells revealed that the 9C4(+) erythroblast population but not the 9C4(-) fraction expressed Ep-CAM mRNA. Peripheral blood CD34(+) cells induced in vitro to differentiate into the erythroid lineage showed strong Ep-CAM expression on days 3 to 7 of culture. The addition of Ep-CAM-specific mAbs 9C4 or KS1/4 to the culture resulted in two- to three-fold up-regulation of Ep-CAM protein expression.Conclusion. mAb 9C4 recognizes Ep-CAM, a molecule expressed in the early steps of erythropoiesis. (C) 2002 International Society for Experimental Hematology. Published by Elsevier Science Inc.