CHROMOSOMAL FLOW CYTOMETRY--DNA AND CENTROMERES

染色体流式细胞术--DNA 和着丝粒

基本信息

批准号：
3272763
负责人：
JOE N LUCAS
金额：
$ 24.6万
依托单位：
UNIVERSITY OF CALIF-LAWRNC LVRMR NAT LAB
依托单位国家：
美国
项目类别：
财政年份：
1978
资助国家：
美国
起止时间：
1978-04-01 至 1990-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3272763
关键词：
DNA centromere chromosome disorders clinical biomedical equipment computer assisted diagnosis computer simulation cytogenetics flow cytometry fluorescence karyotype

项目摘要

Two techniques are available for high resolution flow cytometry of chromosomes in suspension: Dual Beam Flow Cytometry and SlitScan Flow Cytometry (SSFCM). Dual beam flow cytometry of chromosomes stained with Hoechst 33258 (Ho) and Chromomycin A3 (CA3) and similar dye pairs allows the separation of most but not all human chromosomes. Slitscan flow cytometry and centromeric index (C1) calculation can distinguish between chromosomes that bind equal amounts of Ho and CA3 but differ in the position of the centromere. Thus, chromosomes that are difficult to separate by dual beam flow cytometry (e.g. the larger human chromosomes and especially 9 through 12) can be distinguished by SSFCM. Further, SSFCM allows detection of morphologically aberrant chromosomes such as dicentrics. We propose in this application to develop a flow sorter that allows combined dual beam/slitscan flow cytometric analysis to facilitate classification and purification of the human chromosomes. At the heart of this instrument will be a digital pulse shape analysis circuit that allows C1 calculation or dicentric chromosome detection within 1ms. This circuit and the mechanical and optical components of the dual beam SSFCM will be installed on an existing prototype LLNL sorter thereby augmenting development of a dual beam slitscan sorter (DBSSS). This instrument will be a powerful tool: 1) for the detection of homogeneously occurring numerical and structural aberrations for improved cytogenetic analysis, 2) for quantification of the frequency of random aberrant chromosomes to facilitate quantification of genetic damage, and 3) for purfication of chromosomes according to shape, DNA content, and DNA base composition for validation of the above assays and for improved chromosome purification by sorting.

两种技术可用于高分辨率流式细胞仪悬浮液中的染色体：双光束流式细胞术和 Slitscan流式细胞仪（SSFCM）。双光束流式细胞术用Hoechst 33258（HO）染色的染色体和铬霉素A3（CA3）和类似的染料对允许大多数但不是全部人类染色体的分离。 Slitscan 流式细胞仪和丝粒指数（C1）计算可以区分结合相等量的HO的染色体和CA3，但在丝粒的位置有所不同。因此，难以通过双光束流很难分离的染色体细胞仪（例如，较大的人类染色体，尤其是9 通过12）可以通过SSFCM区分。此外，SSFCM 允许检测形态上异常的染色体此类染色体作为dicentrics。我们在此应用中建议开发流程允许组合双光束/狭缝流式细胞仪的分类器分析以促进人类的分类和净化染色体。该乐器的核心将是数字脉冲形状分析电路允许C1计算或二分之一 1ms内的染色体检测。这个电路和双光束SSFCM的机械和光学组件将因此安装在现有的原型llnl划线器上增强双光束狭缝分层器（DBSSS）的开发。该仪器将是一个强大的工具：1）检测同质发生的数值和结构畸变改进的细胞遗传学分析，2）定量随机异常染色体的频率促进量化遗传损害，3）用于纯化根据形状，DNA含量和DNA碱的染色体用于验证上述测定和改进的组成通过排序纯化染色体纯化。