DETECTING AND PHENOTYPING MIXED POPULATIONS OF HIV INFECTED CELLS

在主要未感染细胞中分离罕见感染细胞 背景（如血液）。 目前的艾滋病毒感染诊断测试 取决于循环病毒抗体或病毒抗原的检测 从受感染的细胞中释放出来 没有许可的分析 其鉴定或定量血液中受感染细胞的数量， 其他组织。 有许多检测病毒或 然而，感染细胞的敏感性极限并不好 确立了习 我们计划开发和表征 免疫荧光测定和基于核酸扩增的测定 用于显微镜载玻片以及荧光激活 细胞分选仪和分析仪（FACStar和FACScan）。 FACS功能允许 筛选统计学上显著数量的细胞，并分类 这种能力允许无菌、克隆分离感染的细胞。 这些 技术将用于1）评估感染细胞的数量 在特定的血液亚群中，2）克隆分离 病毒，和3）在体外实验中定位感染位点。 我们 正处于该项目的早期阶段，并已1）开始准备， 验证用于灵敏度测定的对照细胞培养物，2）开始 评价DTTD可用的HIV阳性血清池以及HIV 单克隆抗体，3）FACSTAR Plus操作的培训人员 和4）开发了血液亚群表型鉴定的方案。

Isolation of rare infected cells in a predominantly uninfected cell background (e.g. blood). Current diagnostic tests for HIV infection depend on detection of circulating viral antibodies or virus antigen which is released from infected cells. There are no licensed assays which identify or quantitate the number of infected cells in blood or other tissues. There are numerous research assays which detect virus or infected cells, however, their limits of sensititvy are not well established. We plan to develop and characterize the sensitivity of immune fluorescence assays and nucleic acid amplification based assays for use on microscope slides as well as on the Fluorescence Activated Cell Sorter and Analyzer (FACStar and FACScan). FACS capabilitiy allows screening of statistically significant numbers of cells, and sort capability allows sterile, clonal isolation of the infected cells. These techniques will be used to 1) evaluate the number of infected cells within specific blood subsets, 2) clonally isolate primary strains of virus, and 3) localize sites of infection in in vitro experiments. We are in the early stages of this project and have 1) begun preparing and validating control cell cultures for sensitivity determinations, 2) begun evaluation of HIV positive serum pools available at DTTD, as well as HIV monoclonal antibodies, 3) trained personnel for FACSTAR Plus operation and 4) developed protocols for blood subset phenotyping.