Morphology Resource Core


Morphology Resource Core


 
The Histology section of the Core provides basic services such as

  • Tissue preparation and embedding.
  • Epoxy resin (Epon/Araldite) embedding, sectioning and staining for light and electron microscopy
  • Glycol methacrylate (GMA Technovit 7100) embedding, sectioning and H&E staining for light microscopy.
  • Paraffin block or frozen tissue sectioning and staining The Core also provides additional services including serial sectioning, special stains and technical expertise with immuno-histochemical staining.

Equipment

  • Leica cryostat CM1850
  • Sorvall Porter-Blum MT-2B Ultra microtomes
  • LKB 7800 Knifemaker
  • Zeiss OpMi-1 operating microscope
  • Leica RM2135 manual rotary microtome
  • Leica RM2255 fully automated rotary microtome
  • Leica SM2000R sliding microtome
  • Leica RM2165


 
The Microscopy section of the core provides

  • Leica SP8 confocal microscope is on a TMC anti-vibration table located in K130A. DM6 CS with motorized scanning stage with objectives HS PL APO 10x/0.40 CS2, 40x/1.30 Oil CS2, 63x/1.40 Oil CS2 and 63x/1.30 glycerol correction CS2 plus DIC condenser and objective prisms. Laser lines include Argon/2 (with 458, 488, 514), 405 nm diode, yellow 561 nm, HeNe orange 594 nm and HeNe red 633 nm. Four HyD high sensitivity hybrid detectors, one PMT and a transmitted light detector. HPZ640 workstation (Windows 7) and Leica LAS X system software (Leica Application Suite X) plus the Huygens (SVI) Deconvolution package.
  • Zeiss LSM 700 laser scanning confocal microscope in K230 on a Zeiss Axio Examiner upright body equipped with DIC optics (viAible and IR), a 10X long-working distance dry objective and a 40X long-working distance water immersion objective, as well as, 10X, 20X and 40X dry objectives and 40X and 63X oil immersion objectives. In addition, the microscope has 4 solid-state lasers (405, 488, 555, and 635 nm), a Halogen lamp, a Mercury lamp with filter cubes for fluorescence imaging, and a 4-channel LED Calibri light source (400, 445, 505, and 625 nm) for fluorescence imaging and for light activation of photoreceptors during physiological experiments. The IR-sensitive digital camera can be used for IR viewing of live retinal cells during electrical recordings. The microscope stage and optical shutters are computer-controlled. Imaging and analysis are controlled by Zeiss’s Zen software run on a Fujitsu computer (Windows 7) with a 32-inch flat screen display. The microscope is mounted on an anti-vibration table. The overall setup also includes stage-mountable tissue chambers with perfusion and temperature control, a Sutter MP-285 micromanipulator, and an AxoPatch 1D patch-clamp amplifier. Call or email to set up a time for training.
  • An inverted Zeiss LSM 5 Pascal (in room K130) customized for time-lapse imaging. It includes DIC optics, 5X, 10X, and 20X dry objectives, 40X, 63X and 100X oil objectives, a 40X air corrective objective, a Zeiss AxioCam HR with high resolution color CCD image capture. This microscope has been outfitted with a full environmental incubation chamber from Solent Scientific, which also includes digital temperature control for the chamber, a digital temperature-controlled sample holder, humidifier and carbon dioxide regulation specifically for long-term imaging of live samples. This system has all of the required light sources, computer and monitor, and it is located in the specially ventilated imaging room on a Kinetic Systems Vibraplane vibration isolation table.
  • Zeiss Axiophot epifluorescent microscope in K220. Single color, multiple color and bright field, dark field, phase contrast, DIC images may be captured using the SPOT Flex digital camera and Spot v 5.2 software attached to 17 inch iMAC. Objectives 10 X Plan Apo, 20 X Plan Neofluar, 40 X Plan Apo/oil, 63 X Plan Apo/oil and 100 X Plan Apo/oil. Filters 41007a Cy3 (narrow) 450-700, 41017 Endow GFP 400-650 and UV DAPI with traditional HBO fluorescence illumination.
  • Nikon Optiphot microscope with 4X, 10X, 20X, 40X and 60X dry objectives plus a 5-person viewing head.
  • For Electron microscopy services, we have successfully used the San Francisco VA Hospital Pathology Cell Imaging Lab for some EM services, and we propose to use this model as follows: the Morphology Core will receive fixed eyes and will postfix in osmium tetroxide, embed in epoxy resin, thick section (1μm) and stain tissue blocks. In consultation with investigators, the Core staff will trim the blocks to nearly the desired area for EM analysis, take light micrographs of the area for final trimming, and then contact the EM staff at the VA Hospital. The blocks and drawings (or photographs) will be sent to the VA unit on the UCSF-VA Shuttle, where the VA staff will do final trimming and ultrathin sectioning. The key personnel at the VA unit are familiar with retina and eye structures, so we have them take survey EMs of different magnifications, which are returned to us electronically. When we need different areas photographed or higher magnifications, we communicate by telephone or Email. The VA facility can also do EM immunostaining and EM negative staining.  The services performed by the EM staff at the VA hospital are recharged to individual grant accounts and are very cost-effective and time-saving.


 
The Ocular Imaging section of the Core provides

  • Slit lamp examination and photography of rodent eyes.
  • Fundus photography and fluorescein angiography of rodents using a Micron III unit.
  • Micron III OCT (optical coherence tomography) of mouse and rat eyes at 2-μm resolution.
  • Bioptigen Envisu R4300 for biometric OCT imaging including axial length, vitreous chamber depth, lens thickness and accessing iridocorneal angles with the new telecentric lens.
  • Celeris rodent ERG with light Guide Electrodes that establish robust electrical and optical connections, monitor impedence and ensure corneal hydration. The electrodes are built into the stimulator, reducing setup time to seconds. Test 5-10 subjects per hour (or more, depending on protocol length). Mouse RGB and UV stimulators. Pattern electroretinogram stimulator OLED microdisplay.
  • In vivo rodent eye injection set up with Zeiss OpMi-1 dissecting microscope with FemtoJet micro-injector and NEPA21 electroporator, Sutter P-97 micropipette puller and Narishige microgrinder.

The slit lamp, Micron III and OCT instruments are located in a Procedure Room (PSB 624) in the Barrier Facility of the Animal Care Facility and is restricted to Core vision investigators. Celeris rodent ERG upon request from Dr. Yvonne Ou.

In Procedure room PSB 633 there is a Bioptigen Envisu R4300, Zeiss OpMi-1 operating microscope, isofluorane anesthesia cart and 2 anesthesia chambers set up for set up for in vivo rodent injections.