Multi-Photon Tunable Laser Scanning Microscope

  • 2 and 3 photon-excited fluorescence microscopy/tomography
  • Second and Third Harmonic Generation (SHG/THG) imaging
  • GaAsP dual-channel emission detectors (LSM) and dual-modality (LSM and WF) epi-fluorescence imaging
  • Wide-field and LSM image overlay
  • Static and dynamic imaging with 8 kHz Resonant-Galvo scanner
  • Dry / dipping / immersion high NA objectives with W.D. 1.2 to 3.0 mm
  • Deep Z-stacking and wide tiling
  • Capable of imaging cells, tissue (live, fixed or cleared), or awake non-primate animal subjects (temporary housing available)
  • NI FPGA-based digital signal processing and acquisition electronics with potential for laser synchronization/enhanced noise rejection
  • Matlab-based open-source acquisition software (Vidrio ScanImage Premium)

New Users

Faculty and students who wish to use the microscope should receive laser safety training and follow the instructions the New Users Guide.

Policy

The microscope is available to trained users on an equal basis. For a comprehensive list of policies (including BSL-2 protocols), please see the Facility SOP.

System Specifications

Microscope frame
  • Sutter Movable Objective Microscope (MOM) head, upright and tiltable from -45 to 90 deg
  • 3D movable objective turret, 22 mm XY and 12.5 mm Z range stepper motors
  • User-replaceable emission filters (25 to 30 mm dia.)
Scanners
  • From 15 Hz ( 512 lines uni-directional ) to 120 Hz ( 128 lines bi-directional ) raw frame rate
  • From 128 to 2048 pixels per line, square or rectangular (0.8x to 32x), with beam blanking
  • Processed multiphoton image rates = 1/5 raw frame rate; max 1 MHz / pixel, with tunable laser output
  • Field of View: from 2.1x1.7 to 0.31x0.25 mm^2 (for 10x to 60x objectives)
Auxiliary capabilities
  • Conventional wide-field epi-fluorescence imaging (with cooled CCD camera and blue or green LEDs excitation,  460-580 nm wide-band)
  • Oblique white light illumination via fiber optic waveguide
  • On-line motion compensation and activity tracking
  • Off-line Point Spread Function deconvolution with Volocity 3D
Laser sources
  • Mid-IR amplified laser source, <100 fs pulses tunable from 1200 to 2000 nm @ 1 MHz
  • Near-IR fiber laser fixed at 1064 nm, <300 fs pulses @ 80 MHz, ~6 W max.
  • Visible fixed at 517 nm, 200 fs pulses @ 1 MHz, ~10W max.
Future upgrades and potential capabilities
  • Upgradable to a second galvo-galvo scanner for multi-region dual-excitation or high SNR imaging capabilities
  • Enhanced optics for deep imaging with orange-red chromophores near 1700nm excitation
  • Up to 1000 um fast-Z objective piezo-translation
  • Simultaneous electrophysiological recording/stimulation and connectivity mapping (with Ephus or WaveSurfer software)
  • Environment control with objective heating, as well as drug & gas perfusion
  • Motorized/manual XY stage for culture sample/animal positioning >12 mm
  • Pulse compressor for imaging or 3D photo-manipulation with tunable (680-1000 nm) or fixed (~1040 nm) ultrafast laser pulses
  • Improved cooled, NIR-responsive or Hybrid detectors for greater sensitivity and time-correlated (FLIM/FCS) imaging methods (4 channels max).

References

Contact Us

Dr. Aleksandr Smirnov
Associate Research Scientist
Institute for Physical Sciences  & Technology (IPST)
University of Maryland, College Park
asmirnov@umd.edu