Core Unit D: Imaging Core (Kinsey) a. Objective The objective of the Imaging Core laboratory is to provide investigators access to the following technologies and resources: Standard optical microscopy technologies: ? Olympus 1X71 inverted microscope ? Optics formats (including bright-field, phase, dark field, DIG, and intensified low-light) ? Single, dual, and triple chromophore indirect immunofluorescence ? Green fluorescent protein (GFP) detection ? Cooled CCD color digital photomicroscopy ? Video microscopy (real time and time-lapse) ? In situ hybridization analysis ? Photomicroscopy on 35 mm film (option if desired) Confocal microscopy technologies: ? Nikon TE2000-U inverted confocal microscope ? Objectives: Long working distance: Nikon 4x Superfluor, 20x PlanFluor, 40x PlanFluor, Oil Immersion: Nikon 20x, and 40x PlanFluor+DIC, 60X PlanApo +DIC ? Lasers: 488nm, 543nm and 633nm ? Transmission detector and 4 PMT detector capable of measuring 4 channels of emitted/transmitted light Confocal image acquisition/analysis supporting capabilities: ? IBM PC for data acquisition ? Dell PC for data analysis ? Nikon ES confocal operating software (1 copy) ? Nikon ES image analysis software (1 copy) ? Metamorph 6.1 image analysis software (2 copies) ? In addition: the confocal microscope has a conventional UV source (Sutter LB/LS17) with digitally operated shutter and filter wheel (Sutter Lambda 10-2) connected to the microscope via fiber optic cable for ratiometric fluorescence imaging (ex. Fura dyes). These images are recorded by a Photometries CoolSnap-ES high speed 14 megapixel camera. Live confocal imaging resources: ? Micromanipulators: Xenoworks digital micromanipulators, (2) permanently mounted ? Nikon: Environmental chamber enclosing microscope stage and optics with digital temperature control ? Picospritzer II pressure microinjection system ? CellTram Vario manual microinjection system ? Prime Tech CT150 piezo impact drill ? Tissue culture incubator Tissue processing and embedding facility As detailed in the project descriptions, below, an increasing number of core users have the need to process tissues for histology. The following equipment will be available in dedicated space in the new BRC on the 3rd Floor where the U54 project labs are also located: ? Tissue Processor: The Tissue Tek 500 by Sakura is available as a bench top unit and we have had personal experience with it as a very reliable and user friendly unit. It will run up to 300 cassettes Contracts for full service are available. ? Embedding Center: The Leica EG 1160 is an ergonomic compact unit;Menu-driven, microprocessor controlled system, 5 languages;Ample storage for paraffin, molds, cassettes and blocks;Large cooling area;Totally programmable unit with automatic on/off function (7-day memory);Paraffin dispensed using a foot switch or paraffin mold handle;All temperature and working time parameters fully programmable with memory and battery back-up. ? Paraffin Pot: The Leica EG 1110 holds 3.8 liters of paraffin and is the lowest price of the three available. The paraffin pot makes cleaning and replacing paraffin from the embedding center much more efficient than melting in the embedding unit itself. ? Microtome: Leica RM2125. This is a very reliable unit that several Center labs have experience with. ? Water bath: TBS FWS 120. This is a very reliable unit that several Center labs have experience with. ? Stainline: Leica ST5010 allows programming up to 15 different staining protocols and will run multiple protocols at the same time. It integrates an oven for slide drying, and minimizes user exposure to reagent fumes. It is the most popular staining unit from Leica, with six in the area. A spectrum of staining protocols is currently used by U54 and other center investigators. Currently, the reagents have to be prepared for each stain run or each lab must devote time and resources to their staining protocols. Digital image analysis: ? Quantitative analysis using microscopy technologies listed above: ? Morphometry ? Object identification, counting, and gray or color intensity quantitation ? Image processing (real time background subtraction, mathematical operations, gamma adjustment, pseudo color generation, etc.) ? Cell migration, counting, distribution, shape analysis ? Quantitative dot-blot, immunoblot, and gel whole band analysis ? Spatial intensity quantitation of GFP, calcium and pH by using single and dual-emission chromophores Digital photographic/imaging/document services: ? Network and local workstation service-based color printing on photographic quality paper ? High resolution digital flat-bed scanning of transparent and opaque films, x-rays, polyacrylamide gels, westerns, photographs, etc. ? Adobe? PhotoShop preparation of digital images ? Optical character recognition (convert scanned documents to text files) ? Image file archiving and network file server ? Assist in preparation of documents, images, etc. for online submissions of manuscripts, grants, etc. Graphic and illustration services The Imaging Core provides researchers access to high quality artistic services for all forms of media ranging from 2D and 3D illustration and animation for Powerpoint presentations, website design, brochure and large format poster design for research conferences, grant applications and journal publications. The Imaging Core staff provides support and training to researchers who utilize the same software. The high end graphics and design packages utilized by the Imaging Core staff include the following: ? Adobe Illustrator cs- Computer drawing and graphic layout software. ? Adobe Photoshop cs- Digital Photo editing software. ? Adobe GoLive cs- Web site design software. ? Adobe InDesign cs- desktop publishing software. ? Adobe Acrobat 7 professional- PDF editing software. ? Microsoft Office: Word, Power point, Excel. ? RhinoCeros 2.0 NURBS- 3 Dimensional graphics modeling for windows. ? Macromedia Flash MX 2004- 2 Dimensional computer graphics animation software. ? Media Cybernetics Gel-Pro 4.0- ID-PAGE gel, northern and western blot gel analysis software. ? Magnafire 2.0 Digital microscopy photography acquisition. Technology training/transfer, quality control and consultations in these areas. The major goals of Imaging Core are to facilitate the research of the users by: ? Providing access to techniques, technologies, and equipment not available in their own laboratory. ? Render cost effective imaging, photomicroscopy and image analysis services at lower cost and higher quality than currently available. ? Maintain a central resource for digital image acquisition and output to reduce user costs and ensure consistent image quality for those techniques performed by the user's laboratory. ? Provide consultation with experts in imaging techniques to enhance experimental design, data collection, and data presentation. ? Continue to provide cutting edge services and support by upgrading existing resources as needs dictate and funds allow.
Showing the most recent 10 out of 16 publications
