FLUORESCENCE IMAGING OF LIVE CELLS AND TISSUES

活细胞和组织的荧光成像

• 批准号: 6291347
• 负责人: SUSETTE C MUELLER
• 金额: $ 22.15万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-15 至 2002-04-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6291347
• 关键词: bioassay; bioimaging /biomedical imaging; biomedical equipment purchase; biotechnology; chick embryo; computer program /software; fluorescence microscopy; image processing; neoplastic cell; tissue /cell culture

This proposal requests the funds to purchase three epi-fluorescence microscope systems. Dynamic cellular imaging affords the possibility of determining temporal relationships between cellular structures and subcellular protein localization. Thus, the first system will be specialized for fast collection of low light fluorescence images of cells cultured on the microscope stage. It will comprise an inverted epi-fluorescence microscope, motorized, encoded stage, a high performance CCD camera, and computer with software. The cells will be transfected with one or two mutant green fluorescence protein (GFP) chimeras linked to kinases, GTP-binding actin-associated proteins, and tubulin. Living cells will also be labeled with fluorophore-conjugated antibodies recognizing extracellular epitopes. Metamorph software will be used to control the motorized stage, camera, shutters, and filter wheels during image collection. This software will also be used to process and analyze the images, and for 3D reconstruction and deconvolution. The second microscope system will be used for stereoscopic visualization of tissues expressing GFP or chimeras of GFP. These experiments will monitor invasion/ metastasis of tumor cells in tissues and protein expression during development in a chicken embryo model. The equipment for this system comprises a CCD camera, epi-fluorescence stereoscope, computer and software. The third microscope is a replacement for a basic fluorescence microscope shared by many users. A core of NIH funded investigators will utilize this instrumentation. The microscope will be housed in the Lombardi Cancer Center Microscopy and Imaging Shared Resource. The Principle Investigator, a cell biologist by training, has been director of this facility since 1991, publishing 18 papers utilizing fluorescence microscopy since that time. She has extensive training in microscopy at both the light and electron microscopy levels. In 1999, the Shared Resource was utilized by 46 investigators representing more than 16 departments in the Medical School and Undergraduate campus. Since May 1999, we have trained over 40 investigators in the use of our new confocal microscope. This microscope is currently at full capacity. In addition to the director, a full time manager/microscopist is on hand to assist users. The Shared Resource is maintained by a combination of Cancer Center support and grant support. Approximately 625 sq. ft. of laboratory space is occupied by this resource and will house the proposed equipment. Users contribute to the resource in the form of user charge back fees. An advisory committee meets annually to oversee fees, policies, and to make recommendations for future growth.

该提案要求资金购买三套落射荧光显微镜系统。动态细胞成像提供了确定细胞结构和亚细胞蛋白质定位之间的时间关系的可能性。因此，第一个系统将专门用于快速收集显微镜载物台上培养的细胞的弱光荧光图像。它将包括倒置落射荧光显微镜、电动编码台、高性能 CCD 相机和带有软件的计算机。这些细胞将被一种或两种与激酶、GTP 结合肌动蛋白相关蛋白和微管蛋白连接的突变型绿色荧光蛋白 (GFP) 嵌合体转染。活细胞也将用识别细胞外表位的荧光团偶联抗体进行标记。 Metamorph 软件将用于在图像采集过程中控制电动载物台、相机、快门和滤光轮。该软件还将用于处理和分析图像，以及 3D 重建和反卷积。第二个显微镜系统将用于表达 GFP 或 GFP 嵌合体的组织的立体可视化。这些实验将监测鸡胚胎模型发育过程中肿瘤细胞在组织中的侵袭/转移和蛋白质表达。该系统的设备包括CCD相机、落射荧光体视镜、计算机和软件。第三种显微镜是许多用户共享的基本荧光显微镜的替代品。 NIH 资助的核心研究人员将使用该仪器。该显微镜将安置在隆巴迪癌症中心显微镜和成像共享资源中。首席研究员是一位受过训练的细胞生物学家，自 1991 年以来一直担任该设施的主任，自那时以来发表了 18 篇利用荧光显微镜的论文。她在光学显微镜和电子显微镜水平上接受过广泛的显微镜培训。 1999 年，代表医学院和本科生校园超过 16 个系的 46 名研究人员使用了共享资源。自 1999 年 5 月以来，我们已经培训了 40 多名研究人员如何使用我们的新型共焦显微镜。该显微镜目前已满负荷。除了主管之外，还有一名全职经理/显微镜师随时为用户提供帮助。共享资源由癌症中心的支持和赠款支持共同维护。该资源占用了大约 625 平方英尺的实验室空间，并将容纳拟议的设备。用户以用户退款的形式贡献资源。咨询委员会每年召开一次会议，监督费用、政策，并为未来的增长提出建议。

项目成果

Tumor suppressor function of Syk in human MCF10A in vitro and normal mouse mammary epithelium in vivo.

Syk 在体外人 MCF10A 和体内正常小鼠乳腺上皮中的肿瘤抑制功能。

• 发表时间: 2009-10-15
• 影响因子: 3.7
• 作者: Sung, You Me;Xu, Xuehua;Sun, Junfeng;Mueller, Duane;Sentissi, Kinza;Johnson, Peter;Urbach, Elana;Seillier;Johnson, Michael D;Mueller, Susette C
• 通讯作者: Mueller, Susette C

A simplified immunoprecipitation method for quantitatively measuring antibody responses in clinical sera samples by using mammalian-produced Renilla luciferase-antigen fusion proteins.

一种简化的免疫沉淀方法，通过使用哺乳动物产生的海肾荧光素酶-抗原融合蛋白来定量测量临床血清样品中的抗体反应。

• 发表时间: 2005
• 影响因子: 3.5
• 作者: Burbelo, Peter D;Goldman, Radoslav;Mattson, Thomas L
• 通讯作者: Mattson, Thomas L

Goa1p of Candida albicans localizes to the mitochondria during stress and is required for mitochondrial function and virulence.

白色念珠菌的 Goa1p 在应激过程中定位于线粒体，是线粒体功能和毒力所必需的。

• 发表时间: 2009-11
• 作者: Bambach, Adrienne;Fernandes, Mariana P;Ghosh, Anup;Kruppa, Michael;Alex, Deepu;Li, Dongmei;Fonzi, William A;Chauhan, Neeraj;Sun, Nuo;Agrellos, Orlando A;Vercesi, Anibal E;Rolfes, Ronda J;Calderone, Richard
• 通讯作者: Calderone, Richard

Simultaneous suppression of epidermal growth factor receptor and c-erbB-2 reverses aneuploidy and malignant phenotype of a human ovarian carcinoma cell line.

同时抑制表皮生长因子受体和 c-erbB-2 可逆转人卵巢癌细胞系的非整倍性和恶性表型。

• 发表时间: 2004-02-01
• 影响因子: 11.2
• 作者: Pack, Svetlana D;Alper, Ozgül M;Stromberg, Kurt;Augustus, Meena;Ozdemirli, Metin;Miermont, Anne M;Klus, Greg;Rusin, Marek;Slack, Rebecca;Hacker, Neville F;Ried, Thomas;Szallasi, Zoltan;Alper, Ozge
• 通讯作者: Alper, Ozge

Tumor-targeting nanodelivery enhances the anticancer activity of a novel quinazolinone analogue.

肿瘤靶向纳米递送增强了新型喹唑啉酮类似物的抗癌活性。

• 发表时间: 2008-03
• 影响因子: 5.7
• 作者: Hwang, Sung Hee;Rait, Antonina;Pirollo, Kathleen F;Zhou, Qi;Yenugonda, Venkata Mahidhar;Chinigo, Gary M;Brown, Milton L;Chang, Esther H
• 通讯作者: Chang, Esther H