Morphology Core
Personnel
| Atul K. Bhan, M.D. | Director |
| Dennis Brown, Ph.D. | Co-Director |
| Ms. Quynh Vu | Research Technician |
| Margaret McLaughlin | Research Technician |
Overview
The Morphology Core facilitates the provision of sophisticated morphologic correlation with molecular studies that would not otherwise be available to many CSIBD investigators. It is an underlying premise of this Core that understanding the pathophysiologic importance of factors and processes identified by molecular biological and other approaches is dependent on the assessment of morphologic changes in mucosa, and more refined analysis of protein localization. This evaluation is especially critical for the study of murine models of IBD in which any manipulation or evaluation of markers/inflammatory mediators must be judged against the yardstick of mucosal inflammation.
Services
The Morphology Core facilities have been made available to all members of the Center on a "by need" or "sign up" basis.
Tissue and Cell Fixation: Conventional Tissue Sectioning
Conventional tissue sections are prepared from tissue fixed in paraformaldehyde and embedded in paraffin. Slides with sections on them are provided to investigators for further manipulation or stained by H & E for routine histological analysis.
Immunofluorescence Staining
Immunofluorescence staining of tissue sections is performed by standard techniques, secondary antibody probes coupled to a variety of fluorophores are used, including FITC, TRITC, Texas Red and CY3.
Confocal Microscopy AND Spinning Disk Confocal Microscopy
In studies where whole cells are studied, or thick tissue pieces need to be examined, it is difficult to obtain a detailed picture of fluorescence distribution within the cell by conventional fluorescence microscopy because of interference from out-of-focus fluorescence from multiple focal planes within the thickness of the sample. Laser confocal microscopy overcomes this problem by optically sectioning samples, so that individual images from any focal plane within a sample can be obtained without any interfering fluorescence from the rest of the sample.
Immunoperoxidase Staining
For the characterization of lymphocytes with monoclonal antibodies, frozen tissue sections or cytosmear preparations are fixed in acetone or PLP. Immunoperoxidase staining of tissue sections is also used to screen monoclonal antibodies generated against cellular components.
Immunogold Staining
High resolution immunogold staining is routinely performed by the Core laboratory. It affords the highest degree of resolution, and quantification is possible by counting gold particles associated with specific cellular structures. Immunogold staining is carried out on ultrathin sections of tissue that have been cut from either resin-embedded blocks, or cut by ultracryomicrotomy at low temperature embedding in lowicryl K4M and HM20, freeze-substitution or ultrathin frozen sections.
Central Antibody and Reagent Bank
The Morphology Core maintains a central real and virtual bank of antibodies of interest to CSIBD investigators. Some of the antibodies currently available for this bank are listed in Immunology Core and include antibodies to human and other species with reactivities to a wide variety of lymphocyte and epithelial cell antigens.
Electron Microscopy
The core makes available a JEOL 1011 electron microscope as well as all auxiliary equipment needed to perform EM services for CSIBD investigators.
Services: Digital Image Processing, Archiving and Slide Making
As mentioned above, there has been a revolution in the science of digital image processing in the past few years, and this area continues to evolve very rapidly. The Morphology Core continues to respond to these technical advances by continually updating its facilities to maintain a state-of-the-art imaging center. In addition to the Laser Scanning and Spinning Disk Confocal digital imaging systems, the following imaging resources are available for CSIBD users.
Education
The Morphology Core personnel are available to train individuals associated with the CSIBD in the use of equipment and in the performance of specific techniques. Many of the techniques used in morphologic or immunocytochemical studies, such as tissue sectioning, require the acquisition of technical skills through prolonged supervision and practice. For groups of CSIBD investigators interested in the application or performance of specific techniques, training services provided by the Core include seminars and workshops organized by Dr. Bhan and Dr. Brown.
