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Dynamic Bioimaging Lab


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Dynamic Bioimaging Lab

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INSTRUMENTATION

Both at UHasselt and KU Leuven, different high-end microscopes are at our disposal, that we can use for carrying out any type of investigations, from detailed single molecule imaging to recording videorate datasets of large biomolecular structures.


Top-notch widefield/TIRF microscope at our disposal at UHasselt-BIOMED. This instrument is equipped with structured illumination for super-resolution SIM experiments, but can also be used for off-the-shelf 3D PALM/STORM. The instrument contains a heating unit and CO2 incubation for live-cell experiments, but also a Faraday cage, micromanipulators and perfusion for combined fluorescence-electrophysiology experiments.

Microscope

  • Zeiss Elyra PS.1
  • different low/high NA and magnification objective lenses
  • temperature and CO2 control
  • axial drift control

Excitation

  • 405/488/594/633-nm CW lasers
  • widefield, total internal reflection and structured illumination
  • framewise alternating excitation

Detection

  • PCO Edge 4.2 CMOS 1280x1280 camera
  • Andor iXon DU-897 512x512 EM-CCD
  • Optosplit module for simultaneous two-color detection, for e.g. smFRET experiments

Applications

  • live-cell, time-lapse and large sample tile scans
  • videorate imaging
  • TIRF-based temporal image correlation spectroscopy (TICS)
  • ~150 nm and 3D superresolution imaging via structured illumination (SIM)
  • down to 20 nm and 3D superresolution imaging via PALM/STORM
  • single-molecule and smFRET
  • combined electrophysiology-fluorescence experiments

Homebuilt dedicated 483/635-nm alternating excitation, dual-color dual-polarization detection confocal microscope. This system allows imaging more than 20 parameters per single molecule, making it the ideal system for subnanometer accurate single-molecule FRET studies. 

Microscope

  • Olympus body
  • UPLSAPO 60xW NA1.2 objective lens

Excitation

  • 483/635-nm picosecond pulsed lasers
  • underfilled excitation (large PSF for single molecule imaging)
  • PIE
  • linear polarization

Detection

  • 75-um pinhole
  • blue side: two Laser components COUNT BLUE detectors
  • red side: two Perkin Elmer APDs

Applications

  • FCS
  • single-molecule FRET 
  • single molecule fluorescence

 

State of the art confocal microscope equipped with multiphoton excitation, FLIM, SHG, spectral detection and automated large sample aqcuisitioning.

Microscope

  • Zeiss LSM880
  • different low/high NA and magnification objective lenses
  • temperature and CO2 control
  • axial drift control

Excitation

  • SpectraPhysics MaiTai DeepSee 100 fs pulsed TiSaff 690-1050 nm
  • 488-nm pulsed laser diode
  • 458/488/514/543/633-nm CW lasers
  • rotatable polarization control at the excitation side

Detection

  • 34-channel combined GaAsP spectral QUASAR and PMT detector
  • 32-element spatially resolved Airyscan detector
  • Big2 polarization-sensitive (non-) & descanned detectors
  • Becker-Hickl hybrid detector
  • Becker-Hickl SPC830 time-correlated single photon counting

Applications

  • confocal laser scanning microscopy
  • live-cell, time-lapse 3D via z-stacking and large sample tile scans
  • two/multiphoton/deep imaging
  • fluorescence lifetime imaging microscopy (FLIM)
  • fluorescence correlation spectroscopy (FCS) and image correlation spectroscopy (ICS)
  • label-free imaging via second harmonic generation (SHG)
  • 1.7x superresolution imaging via Airyscan

 

Homebuilt multicolor FLIM microscope

 Microscope

  • Olympus body
  • UPLSAPO 60xW NA1.2 objective lens
  • axial drift control
  • Till Photonics Yanus IV galvo
  • widefield LED triband epifluorescence
  • xenon lamp for transmission

Excitation

  • 405/485/560/635-nm picosecond pulsed quadband
  • 440/510/560/635-nm picosecond pulsed quadband
  • focused overfilled excitation (confocal)
  • PIE/nsALEX
  • linearly polarized

Detection

  • 1-Airy unit pinhole
  • polarization sensitive
  • Picoquant tauSPAD
  • Picoqant PMA hybrid
  • Picoquant Hydraharp 400 4-channel TCSPC device

Applications

  • confocal imaging
  • time-resolved fluorescence (ps - µs lifetimes)
  • FLIM and FLIM-FRET
  • time-resolved anisotropy
  • FCS, PIE-FCCS, ICS, RICS, PIE-ccRICS, tICS...
  • (STED via available phase mask)
  • single-molecule fluorescence

The lab is embedded in two top-notch imaging facilities (Advanced Optical Microscopy Centre at UHasselt and the Molecular Imaging and Photonics division at KU Leuven), so many optical microscopy methods are readily available, such as STED, light sheet, SOFI, correlative light-EM.