Department of Pathology, College of Medicine and Medical Research Institute, Chungbuk National University, Cheongju 361-763, Korea.
Immune Netw. 2010 Dec;10(6):239-46. doi: 10.4110/in.2010.10.6.239. Epub 2010 Dec 31.
Monoclonal antibodies (mAbs) recognizing Class III epitope of CD34 are essential for flow cytometric diagnosis of leukemia.
27H2 mAb was developed from a mouse alternatively immunized with human acute leukemia cell lines, KG1 and Molm-1. Using flow cytometric analysis of various leukemic cell lines and peripheral blood, immunohistochemical study of frozen tonsil, we characterized 27H2 mAb. Antigen immunoprecipitated with 27H2 mAb immunobloted with anti-CD34 mAb. A case of bone marrow sample of acute lymphoblastic leukemia (ALL) patient was obtained at CBNU Hospital. For epitope identification enzyme treatment with neuraminidase and O-sialoglycoprotein endopeptidase (OSGE) and blocking assay with known classIII mAb (HPCA-2) were done.
Only KG1 and Molm-1 revealed positive immunoreactivity. Immunohistochemical staining disclosed strong membranous immunoreactivity on high endothelial venules. Antigen immunoprecipitated by 27H2 mAb showed approximately 100 kDa sized band immunoblotted with anti-CD34 under non-reducing conditions. Epitope recognized by 27H2 mAb disclosed resistancy to both neuraminidase and OSGE treatment and completely blocked with known class III mAb preincubation. CD34 positive leukemic cells in BM of pre B cell ALL patient detected by FITC-conjugated 27H2 and HPCA-2 were identified with similar sensitivity.
A novel murine mAb recognizing class III epitope of human CD34 with high affinity, which is useful for flow cytometric diagnosis of leukemia, was developed.
识别 CD34 类 III 表位的单克隆抗体 (mAb) 对于流式细胞术诊断白血病至关重要。
27H2 mAb 是从小鼠中开发的，该小鼠通过人急性白血病细胞系 KG1 和 Molm-1 进行替代性免疫。通过对各种白血病细胞系和外周血进行流式细胞分析，对冷冻扁桃体进行免疫组织化学研究，我们对 27H2 mAb 进行了表征。用 27H2 mAb 免疫沉淀的抗原用抗 CD34 mAb 免疫印迹。在 CBNU 医院获得了一名急性淋巴细胞白血病 (ALL) 患者的骨髓样本。为了鉴定表位，用神经氨酸酶和 O-唾液糖蛋白内肽酶 (OSGE) 进行酶处理，并使用已知的类 III mAb (HPCA-2) 进行阻断试验。
只有 KG1 和 Molm-1 显示出阳性免疫反应性。免疫组织化学染色显示在上皮细胞血管上有强烈的膜免疫反应性。用 27H2 mAb 免疫沉淀的抗原在非还原条件下用抗 CD34 显示约 100 kDa 大小的条带。27H2 mAb 识别的表位对神经氨酸酶和 OSGE 处理具有抗性，并且在用已知的类 III mAb 预孵育时完全被阻断。用 FITC 缀合的 27H2 和 HPCA-2 检测到前 B 细胞 ALL 患者 BM 中的 CD34 阳性白血病细胞具有相似的敏感性。
开发了一种新型的高亲和力识别人 CD34 类 III 表位的鼠单克隆抗体，可用于流式细胞术诊断白血病。
