Imaging Facility

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Confocal Microscopy

Leica SP1 Inverted Confocal Microscope (located in HSRB)

SP1 Inverted Confocal Microscope Training Document (PDF, 13 pgs)

Microscope

  • FITC, Rhodamine and DAPI filter sets on a Leica DM IRBE inverted microscope
  • Optics for differential interference contrast (Nomarski)
  • Objectives – 10x immersion, 20x oil, 40x oil, 63x water, 100x oil
  • Harvard Apparatus temperature controller for open perfusion micro-incubator for 35mm culture dishes
  • Can accommodate slides or culture dishes

Lasers

  • Titanium-sapphire multiphoton
  • Argon with a 488 nm excitation line
  • Solid state diode with a 561 nm excitation line
  • Helium-Neon with a 633 nm excitation line
  • Spectrophotometer for continuous selectable wavelength bands
  • Scans five channels simultaneously or sequentially
  • Optical sectioning capability

Computer System

  • Windows XP running on a 2.70 GHZ Pentium processor w/ 95 GB hard drive, 1.0 GB RAM
  • Leica software stores images as tif files, which can be copied to a CD or USB flash drive
  • Workstation available with Leica Lite software

Leica TSP SP5 Confocal with Environmental Chamber (located in BBSRB)

  • SP5 Inverted Confocal Microscope Training Manual (PDF, 12 pgs)
  • Environmental Chamber – Completely encloses the microscope, allowing control of temperature, humidity and CO2
  • Motorized Stage – For precise movement in x, z and y. Can accommodate slides or culture dishes with coverslip inserts
  • Differential Interference Contrast Optics (Nomarski) – Brightfield
  • Objectives – 10x, 20x oil, 40x oil, 63x oil, 63x glycerin, 63x water, 100x oil
  • Lasers – 405 diode for UV, argon 488, HeNe 543, HeNe 594, HeNe 633
  • Acousto Optical Beam Splitter (AOBS) – A switching valve for light, which is tunable to channel any laser line onto the sample and simultaneously transmits the emitted light to the detector, eliminating the need for dichroic mirrors, yielding improved image quality and low bleaching
  • Scan Formats – x, y, z, t, l
  • Image Formats – 64 x 64 pixels to 4096 x 4096 pixels, in square or rectangular formats
  • Electronic Zoom
  • Tandem Scanner – Conventional scanner for classical morphology, and a resonant scanner for fast acquisition at 16,000 lines per second or 200 frames per second
  • Optical Field Rotation – Allows rotation of the image 180°
  • Spectral Imaging detection – Includes five confocal channels; allows freely adjustable positioning and bandwidth of the detection range

    Learn more about the AOBS Confocal Microscope.

Leica TCS SP5 Inverted Confocal with Multiphoton Laser (located in BBSRB)


  • Multiphoton Laser – UV range for thick samples or samples in water
  • Visable Light Lasers – Argon with excitation lines 458nm, 476nm, 488 nm, 514nm, and HeNe excitation lines 543nm and 633nm
  • Transmitted Light detector – Differential Interference Contrast (Nomarski) and brightfield
  • Objectives – 10x, 20x oil, 40x oil,  63x glycerin, 100x oil
  • Scan Formats – x, y, z, t, l        
  • Image Formats – 64 x 64 pixels to 8192 x 8192 pixels, in square or rectangular  formats
  • Electronic Zoom
  • Optical Field Rotation – Allows rotation of the image 200°
  • Spectral Imaging detection – Includes five confocal channels; allows freely adjustable positioning and bandwidth of the detection range.
  • Fluorescence Lifetime Imaging (FLIM) – Used to determine cell parameters such as pH value, ion concentration or oxygen saturation and  protein interaction
  • Mark & Find – Defines the current position of the sample and recalls it later from a list of previously stored positions; can mark multiple cells for recording. It can be combined with time series and z scans so dynamic processes can be recorded in a large sample at multiple positions over time, each with an independent z-stack.
  • Tile Scan – Automatically assembles an image from multiple smaller images. Large specimens bigger than the field of view can be viewed as a whole using tile scan.
  • Spectral Separation – Useful for dyes whose emission ranges are close together or partially overlapping
  • Image Acquistion – Download as tif images and save via  NeroStartSmart software to CDs, DVDs, USB flash drives, portable external hard drives