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

核心——自动化细胞计数

• 批准号: 6347265
• 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: 2000-08-30 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6347265
• 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.

自动细胞仪和细胞分散实验室/共聚焦显微镜以及 图像分析设施为研究人员提供了细胞分析 医学博士安德森癌症中心的同行评审补助金。该设施发展 并在单细胞分析中提供尖端技术。细胞 已经建立了表型，增殖和凋亡测定法，并且 根据需要修改多参数分析。免疫表型分析 与凋亡相关的细胞内蛋白质结合 （Bcl-2，Bag-1，Bcl-XL，P53，RB和FAS），增殖和膜脂质 不对称。细胞抗原的定量允许确定 每个细胞的抗体结合能力。非常罕见的事件和祖细胞 通过三重射线检测并分离了亚群 激发/八参数荧光激活的细胞分选（FACS） 随后通过分子细胞遗传学和其他分子进行分析 技术。获取高速细胞分选项和激光扫描 显微镜将提供最新的隔离和分析。激光 共聚焦显微镜已被广泛使用，因此获取 需要第二种仪器。通过使用电荷耦合设备（CCD） - 基于图像分析系统，一种完美捕获图像捕获的方法 开发了注册并已广泛用于分子 细胞遗传学荧光原位杂交（FISH）和比较 基因组杂交（CGH）。鱼也与 凋亡测定法检测正常和白血病细胞中凋亡。这 最小残留疾病细胞的数量，表型和增殖 可以在鱼类分析的情况下确定适合鱼类分析的异常 在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。该设施已为26名调查人员提供了同行的调查员 审查了使用激光共聚焦显微镜为1,797的赠款 去年FACS设施为1,970小时，比过去2次增长33％ 年。核心不断开发了适合该方法的新方法 用户不断发展的需求。