CONTINUOUS MAGNETIC CELL SORTING

连续磁性细胞分选

• 批准号: 2103509
• 负责人: Maciej Zborowski
• 金额: $ 14.18万
• 依托单位: CLEVELAND CLINIC FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-02-18 至 1997-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2103509
• 关键词: CD antigens; atomic absorption spectrometry; avidin; biomedical equipment development; biotin; bone marrow purging; enzyme linked immunosorbent assay; flow cytometry; fluidity; fluorescent dye /probe; human subject; isothiocyanates; method development; monoclonal antibody; nuclear magnetic resonance spectroscopy; phycobilin; transmission electron microscopy

Separation of cells based on their function or surface properties is a basis of many techniques used in cell biology, cancer therapy and in biotechnology. While immunomagnetic cell separation has been utilized in all these areas, it has been hampered by the relatively large size of the paramagnetic label as compared to the target cell and by the relatively crude devices used for separation. This study will have two primary goals: the development of a novel magnetic cell label, and the analysis and design of a continuous, magnetic cell sorter using this label. This new magnetic label consists of ferritin combined to monoclonal antibodies against a particular surface antigen on the target cell. Ferritin is an iron storage protein in mammals. The properties of ferritin labels will be characterized using a number of techniques including: nuclear magnetic resonance spectroscopy, atomic absorption spectroscopy, and transmission electron microscopy. A unique sandwich labeling technique combining fluorescent and magnetic label will be used in combination with flow cytometry to characterize the cell-label binding. Human peripheral lymphocytes and human breast carcinoma derived cells will be used in this study, in combination with monoclonal antibodies against the specific markers: CD3, CD4, CD8, CD16, CD34 and Ber-EP4. These markers identify cells important for the bone marrow transplantation therapy: T cell, helper/inducer, cytotoxic/suppressor, natural killer, stem and breast carcinoma cells, respectively. Due to the complexity involved, the analysis and design of a continuous, magnetic cell sorter will require the use of 3-Dimensional Particle Image Velocimetry (3-D PIV). 3-D PIV, developed at the Ohio State University, the proposed subcontract site of this study, will allow the determination of 3-D velocity vectors of labeled and unlabeled cells in high magnetic fields. The experimental analysis, combined with the relevant equations of motion, will provide information on the system at a micron scale. With this information an optimized cell sorter will be developed. In addition, it will be possible to design more advanced sorters, such as multistage system, in which very high purity or very high selectivity can be obtained in a relatively short time. The ultimate goal is to develop a continuous, magnetic cell sorter for large-scale separation for bone marrow transplantation therapies. Such a sorter will be applicable to rapid, large-scale T cell depletion, cancer cell purging and hematopoietic stem cell enrichment.

基于细胞的功能或表面性质的细胞分离是一种有效的方法。 细胞生物学、癌症治疗和癌症治疗中使用的许多技术的基础。 生物技术.虽然免疫磁性细胞分离已经用于 所有这些领域，它一直受到相对较大的规模， 与靶细胞相比， 用于分离的粗制装置。这项研究将有两个主要目标： 新型磁性细胞标记的开发，以及分析和 使用该标签设计连续的磁性细胞分选机。这个新 磁性标记由铁蛋白与单克隆抗体结合组成 针对靶细胞上的特定表面抗原。铁蛋白是一种 哺乳动物的铁储存蛋白。 铁蛋白标记物的性质将使用多种表征方法来表征。 技术包括：核磁共振光谱，原子 吸收光谱和透射电子显微镜。一个独特 结合荧光和磁性标记的夹心标记技术将 与流式细胞术结合使用以表征细胞标记 约束力人外周血淋巴细胞和人乳腺癌细胞 本研究中将使用细胞，与单克隆 针对特异性标志物的抗体：CD 3、CD 4、CD 8、CD 16、CD 34和 Ber-EP4.这些标记物识别对骨髓重要的细胞 移植治疗：T细胞，辅助/诱导，细胞毒性/抑制， 自然杀伤细胞、干细胞和乳腺癌细胞。 由于所涉及的复杂性，连续的、 磁细胞分选仪将需要使用三维粒子图像 速度测量（3-D PIV）。三维粒子图像测速技术是由俄亥俄州州立大学开发的， 本研究的拟议研究中心，将允许确定 高磁场中标记和未标记细胞的三维速度矢量 领域的通过实验分析，结合相关方程， 运动，将提供微米级的系统信息。与 将开发优化的细胞分选器。此外，本发明还提供了一种方法， 将有可能设计更先进的分拣机，如多级分拣机， 可以获得非常高纯度或非常高选择性的体系 在相对较短的时间内。 最终目标是开发一种连续的磁性细胞分选仪， 用于骨髓移植治疗的大规模分离。这样的 分选仪将适用于快速、大规模的T细胞去除、癌症 细胞净化和造血干细胞富集。