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The Cell and Tissue Imaging (CTI) Center represents the merger of two previous Shared Resources, the Scientific Imaging Shared Resource and the Cell Microinjection and Live Cell Imaging Shared Resource. The CTI is a centralized, highly specialized shared resource available to all St. Jude investigators. The CTI provides expertise in transmission electron microscopy, confocal laser scanning microscopy, multiphoton microscopy, image analysis, cell microinjection, and live cell imaging.
The Cell and Tissue Imaging Center comprises two divisions: the Electron Microscopy Facility and the Light Microscopy Facility.
The Electron Microscopy Facility of the Cell and Tissue Imaging Center is a highly specialized resource utilizing advanced techniques in electron microscopy imaging. The newly renovated facility encompasses approximately 1800 square feet on the Plaza Level of the Danny Thomas Research Tower. A FEI Tecnai G² F20-TWIN transmission electron microscope with a Field Emission Gun and capabilities up to 200 kV has been added to the facility. This scope gives researchers the ability to do automated tomography, low dose exposure, fast spot scan, and photomontage. It is equipped with an Eagle camera that is sensitive enough for cryo TEM. This system is designed to be easily upgraded to meet changing technological needs. The Jeol 1200 EX II electron microscope has also been upgraded with a new 11 megapixel AMT camera. The facility also has several new state of the art pieces of equipment to allow sophisticated procedures such as freeze substitution and cryo immuno. Automated equipment has also been added to enhance the workflow.
The following staff members are available to assist with the hospital’s imaging needs: Sharon Frase, Director of Electron Microscopy, Linda Horner, Ritch Gursky, and Randall Wakefield.
Scientists have long sought to better understand the diverse array of biochemical processes occurring within cells, including those which govern growth, division, survival and death. Such scientific exploration is frequently advanced by the direct observation of cells and tissues. The light microscopy division of the Cell and Tissue Imaging Center employs high end imaging instrumentation, which enables St Jude researchers to utilize a wide variety of cutting edge imaging techniques to peer inside cells and explore their complex structure and function. Such information is paramount to the understanding of the cell and is perhaps most important when cellular processes break down, causing catastrophic diseases such as cancer.
One commonly used method to visualize proteins in cells is to exploit the highly specific interaction between antibodies and antigens to fluorescently label proteins of interest. Using confocal laser scanning microscopy (CLSM), a technique which provides optical sectioning capabilities as well as enhanced signal to noise over widefield imaging, several proteins can be simultaneously localized to various organelles and subcellular compartments. While this method is extremely robust, antibody staining is generally only useful in fixed cells and tissues, limiting the dynamic information that can be obtained.
Recent advances in fluorescent protein technology have greatly expanded scientists' ability to visualize proteins in living cells. Beginning with the discovery of green fluorescent protein (GFP) in the jellyfish Aequorea victoria in the early 1960's, scientists have developed fluorescent proteins in an astounding array of colors that can be fused to virtually any protein in the cell, allowing complex cellular processes to be observed in real time. Spinning disk confocal microscopy (SDCM) is an ideal method for imaging fluorescently labeled proteins in living cells and tissues. Utilizing high sensitivity CCD cameras as detectors, spinning disk confocal imaging minimizes photobleaching and phototoxicity. Combined with incubator enclosures which maintain physiological levels of CO2, humidity and temperature directly on the microscope stage, cells and tissues can be imaged for hours or even days as they develop, divide or undergo programmed cells death, to name just a few examples.
Imaging deep within thick samples, either live or fixed, is also possible. Multiphoton imaging uses packets of two photons of long wavelength light (infra-red range) delivered as femtosecond pulses to excite the fluorophore. This is advantageous because excitation is limited to a small spot where the laser is focused, avoiding photodamage outside of the focal plane. Another advantage is that the long wavelength light can penetrate deeper into specimens, allowing imaging up to several hundred microns into the sample.
Another important area of live cell imaging involves studying how proteins behave and interact with each other within cells, often at the level of one or only a few molecules at a time. Techniques for studying protein-protein interactions and molecular dynamics (such as trafficking and diffusion) include FLIM, FRET, FRAP, and FCS. Subresolution techniques such as STORM and PALM allow structures as small as 20 nm to be visualized, more effectively bridging the gap between light and electron microscopy than ever possible before. TIRF, which provides a thin optical section at the specimen-coverslip interface, offers a large S/N advantage for studying events at membrane surfaces such as protein or vesicle trafficking to the plasma membrane.
The following staff members comprise the Light Microscopy Facility: Victoria Frohlich, PhD, Yannan Ouyang, PhD, Jennifer Peters, PhD, and Jamshid Temirov, PhD.