FLOW CYTOMETER

描述（由申请人提供）：美国国立卫生研究院资助的田纳西大学健康科学中心的研究人员目前迫切需要一种具有至少五个荧光通道检测、紫外激光激发和自动采样功能的流式细胞仪。此申请是为了获得购买Becton-Dickinson LSR流式细胞仪的资金，该流式细胞仪非常适合满足这些需求。目前有几台流式细胞仪在UTHSC校园运行。这些流式细胞仪都没有配备紫外激光器。目前校园里只有一台仪器能够检测五个荧光通道;然而，该仪器专用于细胞分选，缺乏一些研究人员所需的必要检测灵敏度。专用于细胞分选的流式细胞仪未配备UV激发激光器。目前可用的所有其他流式细胞仪仅限于四个荧光通道，并且在此应用中研究者无法访问或以接近最大容量操作。自60年代末和70年代初问世以来，流式细胞术已成为几乎所有涉及细胞和细胞功能分析的研究的支柱。细胞膜标记物、分化标记物或CD抗原的过多的单克隆抗体，以及细胞质成分如内体和溶酶体蛋白的探针使得流式细胞术成为更大的研究资源。荧光染料，如Indo-1，在结合Ca 2+后改变其发射光谱，为分析通过细胞表面受体的信号传递提供了关键工具。研究者在本申请中使用流式细胞术包括检测巨噬细胞摄入的细菌、检测病毒与细胞表面受体的结合、对淋巴细胞上的细胞表面标志物进行多参数分析、定量Ca 2+动员以测量选定细胞群中的信号传导、使用多种GFP测量基因表达以及使用荧光标志物检测膜融合。

DESCRIPTION (provided by applicant): NIH funded researchers at The University of Tennessee Health Science Center currently have critical need for a flow cytometer with a minimum of five fluorescence channel detection, UV laser excitation, and autosampling capability. This application is for funds to purchase the Becton-Dickinson LSR flow cytometer, which is ideally suited to meet those demands. There are currently several flow cytometers in operation on the UTHSC campus. None of those flow cytometers is equipped with a UV laser. Only one instrument currently on the campus is capable of detecting five fluorescent channels; however, that instrument is dedicated for cell sorting and lacks the necessary detection sensitivity required for some investigators. The flow cytometer dedicated for cell sorting is not equipped with a UV excitation laser. All of the other flow cytometers currently available are limited to four fluorescent channels and are either inaccessible to the investigators in this application or are operating at near maximum capacity. Since its inception in the late sixties and seventies, flow cytometry has become a mainstay for almost all research that involves the analysis of cells and cell functions. Monoclonal antibodies to the plethora of cell membrane markers, differentiation markers or CD antigens, and probes for cytoplasmic constituents such as endosomes and lysosomal proteins have made flow cytometry an even greater research resource. Fluorescent dyes such as Indo-1 that changes its emission spectrum upon binding Ca2+ provide critical tools to analyze signaling through cell-surface receptors. The uses of flow cytometry by the investigators in this application include detection of macrophage ingested bacteria, detection of viral binding to cell-surface receptors, multiparameter analysis of cell surface markers on lymphocytes, quantification of Ca2+ mobilization to measure signaling in selected cell populations, measurement of gene expression using multiple GFP, and detection of membrane fusion using fluorescent markers.