Microscopy

The MCIC microscopy laboratory is fully equipped to conduct biological research at the subcellular and ultrastuctural level. We house light and confocal microscopes, a transmission electron microscopes and two scanning electron microscopes. We offer a variety of sample preparation techniques and work closely with investigators to optimize the various experimental procedures. The MCIC personnel provides assistance and/or individual training in the use of the light and electron microscopes, and technical support for electron microscopy sample preparation. In collaborative projects, the MCIC staff helps design light/EM microscopy experiments and assists with the interpretation of the data. The MCIC staff also trains investigators who wish to learn microscopy techniques and conduct their own work (sample preparation and embedding for light or electron microscopy, thin and ultra-thin sectioning, etc.).


To use microscopy services or start a microscopy project:


Light Microscopy
Instrumentation. Several dissecting and light microscopes are available to users. The epifluorescence inverted Leica DM IRB microscope is equiped with the Leica DFC700T digital camera. The microscope has the following filters: (1) UV (Chroma A: BP 340-380/400/LP425), (2) broad band FITC or GFP (Chroma I3: BP450-490/510/LP515, (3) narrow band FITC or GFP (Chroma +L5: 480/40/505/BP527/30, (4) rhodamine (Chroma N2.1: BP515-560/580/LP590).The Leica S6D dissecting microscope is also equipped with the MC170HD digital camera and the NightSea fluorescence system.
Light microscopy auxiliary equipment includes two microtomes for parafin sectioning and a Leica TP1020 tissue processor for sample preparation.
Services. We train users in microscope usage and sample preparation techniques. We also offer sample preparation as a service.

Confocal Microscopy
Instrumentation. The center has a Leica TCS SP6 confocal scanning microscope. in this "SP" model a specrophotometer replaces emission filters giving ultimate flexibility for "filter-design". This feature is particularly advantegeous for difficult dye combinations, or for plant tissue, where autofluorescence may be a problem. AOBS In addition the Acousto Optical Bream Splitter(AOBS) delivers bright, noise-free images with minimal photo damage at high speed scans. This microscope has four excitation lasers that cover excitation ranges of the most commonly used fluorescent dyes: (1) UV (405nm), (2) Argon-blue (488nm and minor lines 458nm and 476nm), (3) DPSS-green (561 nm), and (4) HeliumNeon- far red (633 nm).
Services. The facility staff provides individual training sessions for confocal microscope users. Users must be trained by the MCIC personnel in order to use the confocal microscope.

Laser Capture Microdissection

Instrumentation. Laser capture microdissection allow to isolate specific cells or parts of tissue from thin microscopic sections. Since this processs does not alter the the selected sample can be then tested for DNA, RNA, protein or metabolites. The center has the Zeiss PALM microbeam instrument (http://microscopy.zeiss.com/microscopy/....), which is designed for collecting samples from paraffin embedded or cryo-sectionned tissue. For cryosections we have the Thermo Scientific Shandon Cryotome SMA equipped with the Leica CryoJane tape transfer system. [back to top]

Transmission Electron Microscopy

Instrumentation. The Hitachi H-7500 is characterized by a new electron optical lens system, which allows the acquisition of high contrast images even at low resolution. It operates in a Windows environment and allows user friendly interactive operation. Its magnifications ranges from 700X to 600.000X, with accelerating voltage between 40kV and 120kV. It's equipped with the SIA-L12C digital camera.
Services include (1) biological sample processing for ultra-thin sectioning for morphology study. For ultrastuctural and morphological studies, the biological specimen is fixed, dehydrated and embedded in plastic blocks, which are then sectioned into app. 80 NM sections. After staining with electron dense stained, the sections are observed at the electron microscope. We use a generic fixation and embedding protocol which produces a good fixation and ultra-structural preservation. If you would like to use a different protocol, you need to talk to us.
(2) Negative staining of thin samples. This is a staining technique that allows visualization of particulate biological samples in suspension, such as virus particles, bacteria, bacteriophages and protein or lipoprotein complexes. An electron dense stain containing heavy ions, such as uranyl acetate or lead citrate, is applied to the to the biological sample in suspension. The stain deposits around the specimen, which is then placed onto formovar carbon coated grids for electron microscope observation.
(3) Immunogold labeling. This is a very powerful technique that provides information on the identity, localization and distribution of macromolecules at an ultrastructural level. The techniques involved in this procedure vary and no single preparative procedure can be generally applied and the best method has to be worked out for each experimental system. [back to top]

Scanning Electron Microscopy

Instrumentation. Our Hitachi S-3500N scanning electron microscope can operate in both conventional high vacuum in the specimen chamber, or at elevated pressure. This feature allows for minimal specimen preparation and allows viewing of hydrated biological samples without the conductive coatings required for conventional scanning electron microscopy. A resolution of 3.0nm is obtainable in high vacuum mode, and a resolution of 4.5nm in variable-pressure (VP) mode. The microscope is PC controlled and allows quality digital image recording and storage. The microscope is also equipped with a Noran X-ray analytical device for dispersive X-ray elemental microanalysis.
MCIC users have also access to the USDA cold field emission Hitachi S-4700 scanning electron microscope. This microscope is also equipped with a Noran X-ray dispersive analysis device. If you would like to use this microscope, you need to contact: Leona Horst, Technical Assistant, USDA-ARS, horst.9@osu.edu
Services. We provide a variety of sample preparation techniques, sample critical point drying and coating with carbon, gold or platinum. [back to top]