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Hyperspectral Imaging Laboratory (LSM 510 META)

CAPABILITIES

The Hyperspectral Imaging Laboratory supports:

  • Separation of up to eight fluorophores in a single sample
  • Fluorescence excitation of CFP, GFP, fluorescein, Fluo-3, YFP, rhodamine, and Cy5
  • Ti:Sapphire multiphoton laser excitation, tunable to wavelengths between 700nm to 900nm, for studies of thick samples and ultraviolet-excited fluorophores, and for focalized photolysis
  • Forster resonance energy transfer (FRET) and anisotropy-FRET studies
  • Automated collection of optical sections, for three-dimensional reconstruction
  • Automated collection of time series, for studies of dynamic processes and fluorescence recovery after photobleaching (FRAP)
  • Lateral resolution of up to 250nm, axial resolution of up to 1 um
  • Laser scanning Nomarski differential interference contrast transmission microscopy

Hardware

Zeiss LSM510 META laser scanning confocal microscope 

  • Axiovert 200 inverted microscope
  • Omnichrome Argon ion laser
        ~25mW of 458nm, 488nm, and 514nm excitation light
  • Zeiss Helium/Neon laser with 2mW of green (543nm) excitation light
  • Zeiss Helium/Neon laser with 6mW of red (633nm) excitation light
  • SpectraPhysics MaiTai Ti:S ultrafast laser with 710-990nm tunable excitation wavelengths, up to 1.5W (average power) for multiphoton applications
  • Objectives: 63x/NA1.4 oil immersion plan-apochromat, 40x/NA 1.3 oil immersion plan-neofluor, 40x/NA1.2 water immersion C-apochromat, 25x/NA0.8 multi-immersion plan-neofluor, and 10x/NA0.45 plan-neofluor.
  • Special optics for Anisotropy-FRET applications
  • DIC Optics
  • Long working distance condensor (NA=0.55)
  • Three Hamamatsu photomultiplier tube detectors
        ~Two conventional side-on PMTs
        ~One 32 channel PMT
  • Dichroic mirrors and filters for simultaneous or sequential observations of AMCA/Indo-1, CFP, fluorescein/fluo-3/GFP, YFP, rhodamine/RFP and Cy5 fluorophores, and for reflection confocal microscopy
  • Transmitted light detector

Location

The Hyperspectral Microscopy Laboratory is located in space provided by the Department of Physiology on the fifth floor of the Bressler Research Building, 655 W. Baltimore St, Room 5-040.

Funding Sources

NSF Grant for the Hyperspectral Confocal Microscope

In September, 2004 the Division of Biological Instrumentation and Resources of the National Science Foundation provided $346,692 to establish the Hyperspectral Confocal Imaging Laboratory. The grant was organized by P.Luther, (Physiology), along with co-PIs B. Alger (Physiology), R. Bloch (Physiology), W. Lederer (UMBI), W. Randall (Pharmacology), M. Schneider (Biochemistry), S. Thompson (Physiology), T. Rogers (Biochemistry), J. Wade (Physiology), and E. Weinmann (Nephrology). The University of Maryland School of Medicine and Department of Physiology contributed a total of $148,582 in matching funds.