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Cellular and Molecular Imaging Core Facility
Purpose
The ability to image and understand the events that occur at a cellular or subcellular level is an important aspect of toxicological investigations. Cellular and molecular imaging are also components of the research in the Center. The imaging component of this Core Facility is equipped with 3 major state-of-the-art instrument platforms that can be used in all of the research themes of MCTEH. The Cellular and Molecular Imaging Core Facility is located in the new research wing at USM. Its principal objective is to provide instrumentation and expertise for imaging of toxicant effects in vivo and in vitro for funded and proposed new research by members of the MCTEH.
The techniques include:
1. Imaging of neurons
2. DNA analysis including cell cycle analysis, DNA index and analysis of aneuploid populations
3. Apoptosis analysis including sub G1, TUNEL, Annexin V, mitochondrial transmembrane potential, and caspases
4. Functional assays for calcium ion concentration and pH, oxidative species and changes in membrane potential
Services
The Cellular and Molecular Imaging Core Facility assists investigators with experimental design, tissue collection, sample preparation and analysis of various biomarkers. The core has state-of-the-art instrumentation and experienced personnel who train members from Center investigators' laboratories in performing assays for the various biomarkers. In most cases, it is expected that the Principal Investigator will send a technician, graduate student or post-doctoral fellow to the Core to be trained in the application of a particular assay. After training is completed, that individual would then perform the assays for the project using the Core equipment at the nominal hourly rate. In some instances, full service may be provided and analysis conducted by the core facility manager at an hourly rate plus an additional fee for technician time.
Management
The present Director of the Cellular and Molecular Imaging Core Facility is Dr. Stephen Pelsue. Dr. Pelsue has extensive experience with flow cytometry and more recently has developed experience with confocal microscopy. Currently, he is overseeing the training and supervision of technical staff and students on those instruments.
Equipment
The Flow Cytometry Technology Suite consists of a BD FACS Calibur system equipped with a dual laser system (488 nm & 635 nm) and 6 PMT detectors (4-color, forward scatter, & side scatter). The FACS Calibur has mechanical sorting capabilities that allow for cell collection at a rate of 300 cells/ second. CellQuest 4.0 (BD Biosciences) and WinList 5.0 (Verity Software House) are available for data analysis and acquisition on a dedicated computer.
The Confocal Microscopy Technology Suite consists of the FV300 laser scanning confocal microscope (LSCM) system attached to an Olympus IX-81 motorized inverted microscope with additional fluorescent illumination mounted on an anti-vibration table. The FV300 LSCM is equipped with three lasers (multi-Ar, HeNe-green, & HeNe-red) capable of generating 5 excitation wavelengths (458 nm, 488 nm, 515 nm, 543 nm, & 633 nm) and three PMT detectors (2 fluorescent & 1 transmitted light). A dedicated computer operates image acquisition and analysis using Fluoview Tiempo software, which allows for x-y-z acquisition for 3D structural analysis and time-lapse acquisition for temporal analysis.
The Calcium Imaging Technology Suite enables us to perform short- and long-term calcium imaging on cultured cells or tissue slices. The setup consists of an Olympus IX-71 inverted microscope with fluorescent illumination mounted on an anti-vibration table, an attached Hamamatsu Orca ER CCD camera and a Vincent Uniblitz shutter wheel and controller. A dedicated computer running IP Labs Image analysis software controls both image acquisition and the shutter.
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