CORE--FLUORESCENCE ACTIVATED CELL SORTING (FACS)

FACS 核心设施为细胞提供细胞分析 该计划项目的研究人员。实验室提供了 并开发了单细胞分析的尖端技术。 本次资助的重点是细胞凋亡，TdT-b-dUTP 检测 细胞凋亡（TUNEL）的建立和修改 结合 DNA 和 BUdR 的多参数分析 测量（细胞周期、倍性）、免疫表型分析 祖细胞和谱系限制细胞群， 与细胞凋亡相关的细胞内蛋白（bcl-2、p53、Rb）和 细胞表面抗原包括 fas 和 MDR1。最近，绑定 膜联蛋白 V 与磷脂酰丝氨酸 (PS) 的结合被认为是一项测试 变化之前膜脂结构的变化 通过 TUNEL 检测正在发生凋亡的细胞。定量 细胞抗原现已可用，使我们能够确定 抗体结合能力 (ABC)。细胞动力学变化可 在输注 IUdR 和 BUdR 以及后续治疗的患者中进行了研究 通过流式细胞术分析。荧光原位杂交 (FISH) 也已与细胞凋亡检测相结合，允许 我们可以区分正常细胞和白血病细胞的凋亡。的 残留白血病细胞的数量、表型和增殖 (MRD) 具有适合 FISH 分析的异常 30,000 名正常人中只有 1 人患有白血病 细胞。该测定可预测缓解持续时间，并将 适用于诱导治疗和骨髓后的患者 移植。

The FACS Core facility provides cellular analysis to the investigators of this program project. The laboratory has provided and developed cutting edge techniques in single cell analysis. With the focus of this grant on apoptosis, the TdT-b-dUTP assay for apoptosis (TUNEL) was established and modified for multiparameter analysis in combination with DNA and BUdR measurements (cell cycle, ploidy), immunophenotype to analyze progenitor cells and lineage restricted cell populations, intracellular proteins related to apoptosis (bcl-2, p53, Rb), and cell surface antigens including fas and MDR1. Recently, binding of Annexin V to phosphatidyl serine (PS) was recognized as a test for changes in the membrane lipid structure that precedes changes detected by TUNEL in cells undergoing apoptosis. Quantitation of cellular antigens is now available allowing us to determine the Antibody Binding Capacity (ABC). Cell kinetic changes can be studied in patients with infusion of IUdR and BUdR and subsequent analysis by flow cytometry. Fluorescence in situ hybridization (FISH) has also been combined with the apoptosis-assays, allowing us to discriminate apoptosis in normal and leukemic cells. The number, phenotype and proliferation of residual leukemic cells (MRD) with abnormalities amenable to FISH analysis can be determined at levels of as few as 1 leukemic in 30,000 normal cells. This assay is predictive of remission duration and will be applied to patients after induction therapy and bone marrow transplantation.