FV-MPE Multiphoton Microscope


RL B.716


  • Multiphoton imaging of thick in vivo and in vitro samples
  • Simultaneous photostimulation or dual-line imaging
  • Long, working-distance objectives optimized for multiphoton
  • High-speed extended IR (1300 nm) multiphoton imaging (30 FPS at 512 x 512, 438 FPS at 512 x 32)
  • 4 non-descanned detectors (2 GaAsP and 2 multi-alkali PMTs)


Internal user $30/hour
Internal service fee $69/hour

Detailed Specifications


  • Olympus IX83 fully motorized upright microscope
  • Twin IR laser coupling optics with full laser enclosure for independent control of two IR lasers
  • Motorized mirror assembly permits user-selectable stimulation + imaging or simultaneous dual-laser line excitation
  • AOM laser attenuation (0%-100%, 0.1% increments) provides user selectable laser intensity adjustment and laser beam blanking to minimize photo-damage
  • Fully-automated IR beam expander optimizes beam diameter to maximize image resolution with any selected objective
  • Deep Focus Mode adjusts beam diameter in accordance with laser scattering sample conditions to facilitate deeper imaging
  • Quadralign 4-axis auto-alignment optics provides both 2 horizontal axes (X & Y) and 2 angular axes (Olympus-exclusive) of automatic laser realignment for precise multicolor imaging and precise co-alignment with two-laser systems
  • Prior z-deck linear-encoded motorized x-y stage with z-height adjustment
  • Possibility for upgrade to carry out in vivo electrophysiology

Tandem Scanners

  • Olympus SIM scanner for simultaneous imaging and photostimulation
  • Olympus-patented SIM scanner provides additional galvanometer scanning unit for region-of-interest stimulation simultaneous to imaging for optogenetic stimulation, uncaging, etc.
  • Wide choice of stimulation modes enable users to select point stimulation, rastered regions-of- interest, or Tornado (spiral) scans for high-efficiency stimulation

Left: Crosstalk occurred after simultaneous excitation o GFP and DSRED with a single wavelength IR pulsed laser (950nm). Right: Two fluorescence proteins are clearly separated after individual excitation of GFP and DS-RED with a dual wavelength IR pulsed laser (950nm/1040 nm).

Light Sources

  • Main scanner laser–Spectra Physics INSIGHT DS+ -OL pulsed IR LASER, tunable from 680 nm to 1300 nm, 120 fs pulse width at specimen plane
  • Stimulation laser–Olympus-specific SPECTRA PHYSICS MAI TAI HP DEEP SEE-OL pulsed IR LASER, tunable from 690 nm to 1040 nm, 100 fs pulse width at specimen plane
  • Olympus HGLGPLS widefield fluorescence illumination light source 130 W

Conventional Fluorescence Filters

  • U-FBVW; blue-violet excitation, 400-440/455/460 LP
  • U-FBNA; blue excitation, 470-495/505/510-550 BP
  • U-FGW; green, 530-550/570/575 LP

Both maximum intensity projection from 23 slices

Deep Focus Mode improves image brightness

Left: Normal image; Right: Deep Focus Mode; Image Image data courtesy of: Urs Zielger and Jose Maria Mateos, Center for Microscopy and Image Analysis, University Zurich. Mouse line L15 kindly provided by Pico Caroni FMI, Basel


  • XLPLN10XSVMP, Olympus ultra-long working distance 10X MPE-optimized multi-immersion objective 60NA, WD 8 mm, correct collar for refractive indices 1.33-1.52
  • XLPLN25XWMP2, Olympus ultra 25x MPE water-immersion objective 1.05 NA, 2 mm WD
  • 5x NA 0.1, WD 20 mm dry (MPLN5X-1-7) and 40x, NA 0.80, WD 3.30 mm water-immersion (LUMPLFLN40XW(F)) objectives on swing nosepiece

Optimized for best live-tissue and intravital multiphoton imaging. Widefield design maximizes fluorescence light capture. Correction collar provides correction for spherical aberration across a wide variety of tissue refractive indices 400nm-1600nm transmission coatings. Chromatically corrected for 700nm to 1300nm IR lasers.


  • AOBS serves as tunable dichroic for all lasers

Third harmonic generation imaging of porcine adipose tissue; Unlabeled porcine adipose tissue was irradiated with femtosecond laser at 1250nm, second harmonic is detected from collagen fibers at 625 nm and third harmonic from lipid interlaces at 416 nm.



  • Includes 2 channels of GaAsP PMTs for maximum sensitivity and 2 channels of multi-alkali PMTs for maximum spectral range
  • Transmitted light detector for Brightfield/DIC Image Acquisition

Olympus GaAsP detectors provide approximately twice the quantum efficiency of standard multi-alkali PMTs. All non-descanned detectors are positioned as close as possible to the specimen and signaling optics have been enlarged to facilitate maximum detection efficiency of scattered fluorescence.

Computer and Software

  • HP Windows 764-bit Workstation and 30” Monitor, 2TB HD, 128 & 256 GB SSDs, 32GB DDR3 RAM
  • FVMPE-RS system software including sequence manager, stage control, multipoint mapping, and data analysis
  • Full-feature acquisition software provides complete control of system components including laser wavelength and intensity attenuation, laser coupling optics, scanners, & PMTs
  • Stage Control Software provides XY motorized stage control, map image acquisition for easy target locating, tiling acquisition, and image stitching
  • Multipoint Mapping Software provides high-speed multipoint stimulation, multipoint acquisition, and optogenetic mapping
  • Sequence Manager allows programming of complex imaging tasks with microsecond precision within imaging tasks and millisecond precision between imaging tasks for up to two weeks
  • Embedded 64-bit 3D rendering tool
  • Olympus cellSens Dimension provided for post-acquisition data analysis


iBiology tutorial by Kurt Thorn on multiphoton microscopy

 Software manual

Module 1: Basic introduction and lightpath

Module 2: Dye, detector, and laser selection

Module 3: Detectors

Module 4: Objectives and image acquisition