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Analytical Cytometry / Image Analysis

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Equipment

Coulter Cytomics FC500 flow cytometers: These are user-friendly analytical instruments with one air-cooled argon laser and one air-cooled helium-neon laser that permit simultaneous measurement of five different fluorescent probes. These instruments are used for routine multiparameter analysis of DNA content, cell cycle, immunofluorescence, apoptosis, green fluorescence protein expression, oxidative metabolism and phagocytosis. Two Cytomics FC500 instruments are currently located on the Busch campus, and staffed by Theresa Choi and Joan Dubois. A third Cytomics FC500 is located at CINJ and is staffed by Dan Medina.

Coulter Epics Altra flow cytometer/cell sorter: This instrument was recently upgraded to include a 5-watt tunable UV laser and an air-cooled argon and helium/neon laser. New optics, a new amplifier, an additional photomultiplier tube, and computer software have increased the sensitivity of the instrument for cell sorting and analysis. The Epics Altra is currently located on the Busch campus, and staffed by Theresa Choi.

Leica laser scanning confocal microscope: This is a laser based confocal system that permits measurements of fluorescently labeled tissue sections, individual cells, or particles adhered to matrices over a wide selection of visible wavelengths. The instrument is equipped with 4 air cooled lasers allowing for excitation wavelengths of 458, 476, 488, 514, 561, 594 and 633 nm. Analysis of immunofluorescence as well as functional and biochemical kinetic studies can be performed on viable cell samples. Intracellular distribution of fluorescence can also be quantified. The instrument has extremely high sensitivity and can differentiate between closely emitting chromophores. In addition, the Leica TCS SP2 features digital resolution of 4096 pixels x4096 pixels X12 bits per channel with 4 detector channels and can measure real time response of time-critical sequences due to its fast scan speed of up to 2800 lines/sec. The Leica TCS SP2 has a full complement of objectives for collecting high-resolution 3-D images. Laser scanning confocal microscopy will offer CINJ members improved resolution and dramatically improved rejection of out-of-focus light in comparison to conventional epifluorescence microscopy. Optical sections of 1 to 5 mm can be collected at depths up to 150 mm into a specimen. The image is acquired by illuminating each pixel in series by raster-scanning an exciting laser beam over the specimen; a confocal aperture is used to collect the light emitted from only the excited pixel. The Leica confocal microscope is currently located on the Busch campus, and staffed by Dr. Carol Gardner.

Xenogen IVIS® imaging system: This instrument provides investigators with the capability to perform real-time in vivo imaging of small animals. The ultra-sensitive CCD cameras, hardware and software allow detection of emitted photons and the creation of images and data files. The technology makes possible the tracking of biological activity in real-time, at the molecular level. This real-time in vivo imaging utilizes luciferase to visualize markers of interest. By imaging the whole animal at many individual time points, researchers can better understand disease pathology, pharmacokinetics, response to therapy, and other contextual or spatial aspects of the processes taking place in the living animal. The Xenogen IVIS is currently located on the Busch campus, and staffed by Dr. Vasanthi Sunil.