NIA/RRB Confocal Imaging Facility

NIA/RRB 共焦成像设备

• 批准号: 7732257
• 负责人: Kevin Becker
• 金额: $ 44.51万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7732257
• 关键词: Arts; Biochemical; Cell physiology; Cells; Collaborations; Confocal Microscopy; Consultations; DNA; DNA Damage; Development; Extramural Activities; Image; Image Analysis; Imaging Techniques; Immunofluorescence Immunologic; Intramural Research; Investigation; Lasers; Life; Methodology; Metric; Microscope; Microscopy; Modeling; Operative Surgical Procedures; Postdoctoral Fellow; Range; Research Personnel; Scanning; Scientist; Score; Services; System; Systems Analysis; Techniques; Time; Training; Update; base; fluorescence microscope; human WNT2 protein; intracellular protein transport; protein distribution; protein localization location; protein transport; reconstruction; submicron

The Confocal Imaging Facility (CIF) of the NIA IRP has been in operation since October 2004. This facility has one older model (Zeiss LSM 410) confocal system with low usage by IRP investigators. The NIA CIF has been managed and overseen with the expansion and successfully integration of a state of the art Zeiss LSM 510 Meta confocal system, updating the Zeiss Axioskop fluorescent microscope MCID/QImage analysis system and using Metamorph/Coolsnap imaging and analysis system. This has resulted in extensive collaboration with intramural scientists at different levels of expertise ranging from principal senior investigators through postdoctoral fellows and intramural research trainees. We have trained over thirty investigators to be independent users of the confocal microscope and have held scores of consultations with NIA researchers to assist with their imaging and image analysis. We have helped users integrate imaging into their experimental systems and in troubleshooting problems. This has resulted in an increase in the use of immunofluorescence and imaging at the IRP, as attested by the widespread use of the CIF last year by over 60 researchers, ranging from post-bacs to senior investigators. The following techniques have been used or introduced at the CIF: (a) Use of confocal microscopy for precise sub-cellular localization and co-localization of proteins; (b) Advanced immunological, biochemical and imaging techniques required for the investigation of intracellular and intranuclear protein trafficking; (c) Development of methodology to cause DNA damage to live cell DNA at the sub-micron level by use of the continuous scanning UV laser of the LSM 510 Meta system; (d)Time-lapse, FRAP and ratio-metric analysis of cellular processes in live cells using the 510 Meta confocal microscope; (e) Volumetric (3D) reconstruction of intracellular protein distribution using confocal or deconvolution techniques.

NIA IRP的共焦成像设备自2004年10月以来一直在运行。该设施有一个较旧的型号(蔡司LSM 410)共焦系统，IRP调查人员使用较少。通过扩展和成功集成最先进的蔡司LSM 510亚共焦系统、更新蔡司Axioskop荧光显微镜MCID/Q图像分析系统以及使用Metamorph/CoolSnap成像和分析系统，对NIA CIF进行了管理和监督。这导致了与不同专业水平的校内科学家的广泛合作，从首席高级调查人员到博士后研究员和校内研究实习生。我们已经培训了30多名研究人员成为共焦显微镜的独立用户，并与NIA的研究人员进行了数十次咨询，以帮助他们进行成像和图像分析。我们已经帮助用户将成像集成到他们的实验系统中，并对问题进行故障排除。这导致了免疫荧光和成像在IRP的使用增加，正如去年60多名研究人员广泛使用CIF所证明的那样，从BACA后到高级研究人员。 在CIF上使用或采用了下列技术：(A)使用共聚焦显微镜对蛋白质进行精确的亚细胞定位和共定位；(B)研究细胞内和核内蛋白质的运输需要先进的免疫学、生化和成像技术；(C)利用LSM 510Meta系统的连续扫描紫外光激光开发方法学，在亚微米级别对活细胞DNA造成DNA损伤；(D)使用510Meta共焦显微镜对活细胞中的细胞过程进行时间推移、FRAP和比率测量分析；(E)使用共聚焦或去卷积技术对细胞内蛋白质分布进行体积(3D)重建。