Gazitt Y, He Y J, Graham-Pole J
Department of Pediatrics Hematology Oncology, University of Florida Medical Center, Gainesville 32610.
J Immunol Methods. 1992 Apr 8;148(1-2):171-8. doi: 10.1016/0022-1759(92)90170-x.
A methodology for rapid isolation of neuroblastoma (NB) cells from marrow with metastatic NB cells was developed using a cocktail of five antibodies and magnetic microspheres coated with secondary antibodies. Cells bound to microspheres were released by brief exposure to chymopapain, followed by repeated culture of released cells in serum supplemented DMEM medium and selection for adherent cells. Using this methodology, over 35 primary cell lines were obtained free of contaminating normal cells. Detailed analyses of over 14 cell lines revealed gross differences in cell phenotype, size, morphology development of neurite processes, and doubling time (40h-80 h). All cell lines expressed the 145 kDa neurofilament (NF) and a few expressed the 200 kDa NF, with very little or no expression of the 68 kDa NF. DNA analyses revealed 80% near diploid cell lines. High expression of the MDR-1 protein was detected in 6/22 of the cell lines tested.
利用五种抗体的混合物以及包被有二抗的磁性微球，开发了一种从含有转移性神经母细胞瘤（NB）细胞的骨髓中快速分离NB细胞的方法。通过短暂暴露于木瓜凝乳蛋白酶来释放结合到微球上的细胞，随后将释放的细胞在补充有血清的DMEM培养基中反复培养，并选择贴壁细胞。使用这种方法，获得了35个以上不含正常细胞污染的原代细胞系。对14个以上细胞系的详细分析揭示了细胞表型、大小、神经突过程的形态发育以及倍增时间（40小时至80小时）的显著差异。所有细胞系均表达145 kDa神经丝（NF），少数表达200 kDa NF，而68 kDa NF的表达极少或不表达。DNA分析显示80%的细胞系接近二倍体。在所测试的22个细胞系中的6个中检测到MDR-1蛋白的高表达。
