Imaging - Animal - Mediator - Histology
Cystic Fibrosis Shared Imaging Core
In BRB 841, we have two microscopes, one inverted and one upright, cameras and computers available to capture images taken. The capabilities of these facilities are: One inverted Zeiss scope, fully computer controlled. One upright Leica Scope, fully computer controlled with
Solent
Scientific incubator and laser tweezers. Cell perfusion equipment (Bioptech, Zeiss) available. Richardson Test Slide (Generation III) available for critical quantitative imaging. Autoquant and Huygens Essential deconvolution software are available. We are quite a creative bunch, so if you have a special or unusual requirement, talk to us- we can probably figure out a way to accommodate you!
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Leica DM6000 Upright Microscope
[Solent Scientific incubator not shown] |
Leica DM6000 Upright Microscope
MetaMorph 7.0 Controlled
Prior motorized stage
Wide-field fluorescence
Bright Field Microscopy
Differential Interference Contrast (DIC)
Phase Contrast
Polarization
Available objectives:
* 1.25x na 0.04 HCX plan Fluotar
* 10x na 0.3 Plan Fluotar phase
* 20x na 0.5 apochromat long working distance U-V-I dipping
* 40x na 0.8 apochromat long working distance U-V-I dipping
* 63x na 0.9 apochromat long working distance U-V-I dipping
* 63x na 1.20 HCX plan apochromat water immersion
Photometrics 512B Cascade 16-bit camera
* Pixels 512 x 512
* 16 x 16 microns pixel size
* on-chip multiplication gain
Point Grey Research ‘Flea’ USB video camera coupled to Optical Insights Dual-View system
* Pixels 1024x768
* 1/3” Sony ICX204 CCD
* 4.65 x 4.65 microns pixel size
* 3.75 to 30 frames per second, direct digital output.
* Dual-view filter sets: FITC/TRITC, SNARF-1, R and B Phycoerythrin
Leica fluorescence filters in automated turret:
A4 (470 nm emission) - I3 (515 nm emission)
L5 (527 nm emission) - TX2 (645 nm emission)
A (Fura-2)
Laser Tweezers
* 0.5 Watt Nd:YAG laser
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Inverted Zeiss Axiovert 200M
MetaMorph 6.2 controlled
Wide-field fluorescence
Bright Field
Differential Interference Contrast (DIC)
Phase Contrast
Available objectives:
* 10x, 20x phase contrast
* 40x long working distance
* 100x and 63x oil immersion
* 63x water immersion
* 1.6x Optovar can be used, increasing magnification
Photometrics Coolsnap HQ digital camera (12 bit depth)
* 1392 x 1040 imaging array
* 6.45 x 6.45 micron pixels (Sony ICX285 CCD chip)
* 8.77 x 6.6 mm imaging area
* Shutter speeds adjustable from 1 microsecond to 10 seconds
Fluorescence filters available:
* DAPI, FITC/TRITC, Texas Red, GFP, YFP, Cy5, GFP/YFP (Fret), Fura 2, Endow GFP, SPQ, JP2
Cell Perfusion Equipment
Zeiss Incubator S-M
Zeiss Tempcontroller 37-2
Zeiss CTI controller 3700 (Heating, CO2)
Bioptechs objective controller
Bioptechs FCS2 controller
Warner Instruments RC-50 two-sided chamber
Switchable 6-reservior temperature controlled culture medium feeder
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Richardson Calibration Slide:
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Data Collection:
Use the Photometrics camera to save complete image stacks or single images directly onto our hard drive. Files can be large- a single full-frame image is approximately 3 MB. Single binning increases resolution and file size, while 2x2 binning is a good trade-off for increased sensitivity (and the file size is significantly smaller). Metamorph allows for complete automated image acquisition of a z- stack, in time series, at multiple locations, and combinations of all three. Image data needs to be removed from the on-line computer in a timely fashion. Files can be electronically moved to the off-line workstation.
Data Analysis:
Once you have 3D stacks of data, we mostly use Autoquant or MetaMorph to process the images and extract interesting information (colocalization, optical density) from the data. We have installed on the Windows off-line workstation
* MetaMorph 4.6
* Autoquant 9.0
* IPLab 3.03
Scion Image
We also have Huygens Essential deconvolution software, version 2.9, on our Linux dual processor workstation. Data can be written to DVD via Roxio.
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Schedule:
Users can directly schedule time for microscope use. Email or call me if you'd like to use the microscope!
Use of the offline workstation is provided free of charge to users of the facility.
If you have questions or would like more information, please contact:
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Animal Core
The Pediatric Animal Core is directed by world-renowned geneticist Dr. Mitchell Drumm and is managed by veterinarian Dr. Anna M. van Heeckeren. Our mission is to provide the best mouse models available by ensuring genetic quality and perfect health, while maintaining the humane care and use of the mice bred, maintained, distributed and used by Core personnel.
Additional services include teaching techniques using mice humanely, such as inducing disease states to model the human condition, substance administration, euthanasia, and blood and tissue collection, or conducting research projects.
Mice and research services are available on a first come, first serve basis to investigators within the Pediatric Department, collaborators of those investigators, as well as investigators outside the University.
Our web site is cfac.cwru.edu, providing more information about our services and fees.
Inflammatory Mediator Core
The Inflammatory Mediator Core laboratory performs assays of inflammatory mediators, cytokines, intracellular second messengers as well as measurement of urea and protein for normalization of results on human and murine BAL samples, culture media, blood samples, and sputum samples.
Common assays include human cytokines IL-6, IL-8, IL-17, GM-CSF, and murine cytokines including MIP-2, KC, TNF-alpha, IL-1beta, and IL-6. Also routinely measured are eicosanoids, cyclic nucleotides, and various murine growth hormones. Essentially all cytokine assays are now performed utilizing the Luminex LabMAP analyzer. The use of the Luminex analyzer allows for the simultaneous analysis of multiple cytokines from individual samples resulting in more accurate direct comparisons of cytokine expression profiles, faster processing of assays, and a lower cost per assay.
For information, contact Tom Kelley (tjk12)
Histology core
The Histology Core operates out of the eighth floor of the
Biomedical
Research
Building
and is staffed by Claudia Garner, under the direction of Dr. Alan Tartakoff, Professor of Pathology. Consultation with Dr. Tartkaoff is often the initial entrée into this core, as he will identify the services needed and the most appropriate means of meeting the need. Electron microscopy is available at reduced cost for investigators working on cystic fibrosis through consultation with Dr. Tartakoff. For histology, this Core will prepare frozen or fixed sections of tissue or cell culture samples, and perform routine stains on them. Antibody staining and special stains are the responsibility of the investigator, but the Core will work with the investigator to obtain the best substrate or to perform counterstains. Blocks will be returned to the investigator. For investigators working on the cystic fibrosis problem, there is no charge for these services. For investigators working on other problems, charges may be rendered depending on the complexity and number of samples.
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