Current Facility

In March 2003, the Indiana Center for Biological Microscopy moved to new facilities, nearly doubling the area of the Center to a 1200 sq ft floor plan, as well as improving space utilization by optimizing the design of each room for each microscope system. Rooms that have been added are for animal surgery and cell culture. The Facility is equipped with the following major equipment:

Confocal Microscope Systems

A Bio-Rad MRC1024 Laser Scanning Confocal/Multiphoton Microscope is equipped with a Krypton-Argon (488, 568, 647 nm excitations) laser for confocal microscopy and a tunable Titanium-Sapphire laser (using a 5W Millennia diode solid state pump laser) for multiphoton microscopy. The system is equipped with 3 detectors in both the epi-illumination and transillumination paths, as well as three channel external detectors for 2-photon imaging.  The system supports confocal imaging of green, red and far-red emitting fluorophores and multi-photon imaging of blue, green and red emitting fluorophores.  The system can also be used to collect bright field and Nomarski images.  The system is mounted on a Nikon TE-200 inverted microscope.

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A Zeiss LSM-510 Meta Confocal/Multiphoton Microscope System equipped with an Argon laser (458, 488, 514nm excitation) and two Helium-Neon Lasers (543 NM and 633 NM excitation) for confocal microscopy, and a tunable Titanium-Sapphire laser (using a 10W Millennia diode solid state pump laser) for multiphoton microscopy.  The epifluorescence can be captured via two conventional PMT detectors or via the new Meta system.  The Meta system is an array of 32 detectors, which permits collection of the total emission spectrum from a fluorescent sample.  This system allows users to configure the system to collect specific emission ranges or, when combined with the linear unmixing software, to deconvolve sample emissions to allow sensitive discrimination of a large number of spectrally overlapping flourophore  Multiphoton microscopy images can be collected either via these detectors or two external detectors.  This system supports confocal imaging of green, red and far-red emitting fluorophores by confocal microscopy, and multi-photon imaging of blue, green and red emitting fluorophores.  The system is mounted on a Zeiss upright microscope.

 

A Zeiss UV LSM-510 Confocal Microscope System is equipped with a UV Argon Laser (351 NM, 364 NM excitation), a visible Argon laser (458, 488nm excitation) and two Helium-Neon Lasers (543 NM and 633 NM excitation). The microscope is equipped with four epifluorescence detectors and one transillumination detector.  This system supports confocal imaging of blue, green, red and far-red emitting fluorophores and can also be used to collect bright-field or Nomarski images.  The system is mounted on a Zeiss Axiovert 100 inverted microscope.

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A Spinning Disk Confocal system equipped with 3 lasers providing excitations at 442 NM, 488nm, 514 NM, 568 NM and 647 NM  This system utilizes a unique spinning disk that simultaneously scans hundreds of spots over a sample, simultaneously collecting images with an Ixon air cooled EMCCD camera (Andor).  This design allows the system to collect images at over 30 frames per second with very low levels of illumination.  The reduced photobleaching and phototoxicity of this system makes it especially suited to live cell imaging.  This system is configured to rapidly collect images of cells expressing CFP and YFP, or labeled with green, red and far-red emitting fluorophores.  It can also collect bright-field and phase contrast images.  The system mounted on a Nikon TE-2000U inverted microscope.

 

Widefield Microscope Systems

A Nikon Eclipse TE200 Inverted Microscope equipped with a Hamamatsu 1394 Orca-ER Cooled CCD Camera and Micro-Manager software. The highly sensitive Hamamatsu Orca-ER camera is ideal for detection of fluorescence in live samples and in samples that have low signal levels. This high performance widefield microscope system is also equipped with Differential Interference Contrast optics (DIC). The inverted microscope stand accomodates slides or culture dishes. Best results are obtained using thin samples, such as well spread single cells. The system is equipped to collect images of blue, green, red and far-red emitting fluorophores.

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A Nikon Diaphot 200 Inverted Microscope equipped with a Diagnostic Instruments SPOT color camera. The limited sensitivity of this system makes it inappropriate for epifluorescence of anything but very bright probes, but the system is optimized for high-resolution Nomarski, DIC and phase contrast imaging. An Eppendorf micromanipulator and microinjector is available to mount on the stage of the microscope making it particularly useful for imaging living cells over time.

 

A Nikon Microphot SA Upright Microscope equipped with a sensitive Diagnostic Instruments SPOT RT Slider color camera capable of capturing moderately low-light level images of blue, green and red emitting fluorophores. This system is very simple to use, and in addition to providing excellent epifluorescence images, is capable of collecting color, bright field and phase contrast images.

 

Auxiliary Equipment available for use with scopes:

(3) Warner DH-35 dish warmers, a PMDI Open Perfusion Micro-incubator, a Warner RC-50 Transepithelial imaging chamber, and(2) Warner OW objective warmers.

Computer Systems

In addition to the scope systems, the facility has a 5.4 terabyte IBM Fileserver and two public workstations. Web and remote file services are available via a failover linux cluster. Interactive 3D (voxel-based) image processing is performed using PCs equipped with 3.2 GHz Xeon processors, FireGL and Radeon-based video boards, and 6 to 8GB of memory. Various other image-oriented programs are also run on the PCs, including Media Cybernetics' AutoDeblur for deconvolution, Universal Imaging's Metamorph for image analysis/processing, NIH's ImageJ, Amira, Adobe Illustrator and Photoshop for image editing, and Premiere for making movies.

Publication-quality images and graphics can be generated on a Kodak 8670 PS dye sublimation photo printer, Xerox Phaser 7300 color laser printer, and an Epson Photo 2200. Most PCs are equipped with CD burners and some offer DVD-R/RW recorders. Core microscopy systems and workstations are connected by gigabit ethernet.