CORE--AUTOMATED CYTOMETRY & CELL SORTER LABORATORY/CONFOCAL MICROSCOPY

核心——自动化细胞计数

• 批准号: 6352698
• 负责人: MICHAEL ANDREEFF
• 金额: $ 25.41万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-07-18 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6352698
• 关键词: apoptosis; bioimaging /biomedical imaging; biomedical facility; cell population study; charge coupled device camera; confocal scanning microscopy; flow cytometry; fluorescent in situ hybridization; human tissue; image processing; neoplastic cell; single cell analysis

The Automated Cytometry and Cell Sorter Laboratory/Confocal Microscopy and Image Analysis Facility provides cellular analysis to investigators with peer-reviewed grants at M.D. Anderson Cancer Center. The facility develops and provides cutting-edge techniques in single-cell analysis. Cell phenotyping, proliferation, and apoptosis assays have been established and modified as needed for multi-parameter analysis. Immunophenotypic analysis was combined with assays of intracellular proteins related to apoptosis (bcl-2, BAG-1, Bcl-Xl, p53, Rb, and fas), proliferation and membrane lipid asymmetry. Quantitation of cellular antigens allow determination of antibody binding capacity per cell. Very rare events and progenitor cell subpopulations have been detected and isolated by three-laser excitation/eight-parameter fluorescence-activated cell sorting (FACS) for subsequent analysis by molecular cytogenetics and other molecular techniques. Acquisition of a high-speed cell sorter and a laser-scanning microscope will provide state-of-the-art isolation and analysis. Laser confocal microscopy has been used extensively, and so the acquisition of a second instrument is necessary. By using a charge coupled-device (CCD)- based image analysis system, a method for image capture in perfect registration was developed and has been used extensively for molecular cytogenetics fluorescence in situ hybridization (FISH) and comparative genomic hybridization (CGH). FISH has also been combined with the apoptosis assay to detect apoptosis in normal and leukemic cells. The number, phenotype, and proliferation of minimal residual disease cells with abnormalities amenable to FISH analysis can be determined at levels of as few as one malignant cell in 30,000 normal cells. Methods to detect transgene expression in cells have been established using beta- galactosidase (beta-gal), nerve growth factor receptor (NGF-R), and green fluorescent protein. The Facility has served 26 investigators with peer- reviewed grants who used the laser confocal microscope for 1,797 and the FACS facility for 1,970 hours last year, a 33% increase over the last 2 years. The Core has continuously developed new methodology to suit the evolving needs of its users.

自动细胞计数和细胞分选实验室/共焦显微镜和 图像分析设施为研究人员提供细胞分析 M.D. 安德森癌症中心经过同行评审的资助。该设施发展 并提供单细胞分析领域的尖端技术。细胞 表型分析、增殖和凋亡分析已经建立并 根据多参数分析的需要进行修改。免疫表型分析 与细胞凋亡相关的细胞内蛋白测定相结合 （bcl-2、BAG-1、Bcl-Xl、p53、Rb 和 fas）、增殖和膜脂 不对称。细胞抗原的定量可以确定 每个细胞的抗体结合能力。非常罕见的事件和祖细胞 已通过三激光检测并分离出亚群 激发/八参数荧光激活细胞分选 (FACS) 随后通过分子细胞遗传学和其他分子分析 技术。购置高速细胞分选仪和激光扫描仪 显微镜将提供最先进的分离和分析。激光 共聚焦显微镜已被广泛使用，因此获取 需要第二台仪器。通过使用电荷耦合器件（CCD）- 基于图像分析系统，一种完美的图像捕捉方法 注册已开发并已广泛用于分子 细胞遗传学荧光原位杂交（FISH）和比较 基因组杂交（CGH）。 FISH 还与 细胞凋亡检测可检测正常细胞和白血病细胞的细胞凋亡。这 微小残留病细胞的数量、表型和增殖 可以在水平上确定适合 FISH 分析的异常 30,000 个正常细胞中只有一个恶性细胞。检测方法 已经使用β-建立了细胞中的转基因表达 半乳糖苷酶（β-gal）、神经生长因子受体（NGF-R）和绿色 荧光蛋白。该机构已为 26 名研究人员提供了服务 审查了使用激光共聚焦显微镜的赠款 1,797 和 FACS 设施去年运行了 1,970 小时，比过去 2 年增加了 33% 年。核心不断开发新的方法来适应 用户不断变化的需求。