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

核心——自动化细胞计数

• 批准号: 6101817
• 负责人: MICHAEL ANDREEFF
• 金额: $ 33.23万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2000-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6101817
• 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仪实验室/共聚焦显微镜和 图像分析工具为调查人员提供细胞分析 安德森癌症中心的同行评议拨款。该设施开发 并提供单细胞分析的尖端技术。细胞 已经建立了表型、增殖和凋亡分析，并 根据需要进行修改，以进行多参数分析。免疫表型分析 与细胞内与细胞凋亡相关的蛋白质的检测相结合 (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 用三束激光探测和分离了亚群 激发/八参数荧光激活细胞分选(FACS) 后续的分子细胞遗传学和其他分子分析 技巧。高速细胞分选机和激光扫描仪的研制 显微镜将提供最先进的分离和分析。激光 共聚焦显微镜得到了广泛的应用，因此获得了 第二个工具是必要的。通过使用电荷耦合器件(CCD)- 一种基于图像分析系统的完美图像捕获方法 配准是发展起来的，并已被广泛应用于分子 细胞遗传学荧光原位杂交及其比较 基因组杂交(CGH)。鱼也已经与 用细胞凋亡率检测正常细胞和白血病细胞的凋亡率。这个 微小残留病细胞的数量、表型和增殖 对于符合FISH分析的异常情况，可以在水平上进行确定 在30,000个正常细胞中只有一个是恶性细胞。要检测的方法 转基因在细胞中的表达已建立在使用β- 半乳糖苷酶(β-GAL)、神经生长因子受体(NGF-R)和绿色 荧光蛋白。该机构已经为26名调查人员提供了同行- 回顾了1797年使用激光共聚焦显微镜的资助金和 FACS设施去年使用了1,970个小时，比过去2年增加了33% 好几年了。核心不断开发新的方法，以适应 不断变化的用户需求。