Lattice Light Sheet Microscope

The Lattice Light Sheet Microscope (LLSM) achieves fast fluorescence imaging with decreased light dose at the sample plane compared to conventional imaging methods. The system generates an optical lattice to create a 400 nm-thickness light sheet to image biological samples over long periods of time and with very fine resolution. This feature of LLSM allows for 4D living cell imaging, where experiments limited to seconds or minutes on other imaging platforms, can be extended to hours or even days.

HL-60 cell migrating through collagen matrix (the video from Science. 2014, 346, 1257998)

RESERVE THIS MICROSCOPE

System specifications

• TEMPORAL RESOLUTION: ∼100 frames/s, ∼1 cell volume/s
• IMAGING TIME: up to 48 hours
• LIGHT SHEET THICKNESS: 0.4µm at 50µm length
• SAMPLE CHAMBER: Medical grade stainless steel with temperature (TEC) & CO₂ (5%) control
• SPECIMEN MOUNTING: Standard, horizontally-oriented 5mm round coverslip
• ANALYSIS WORKSTATION: Dual 10-Core Xeon 2.4GHz processors, 256GB RAM, NVIDIA Quadro P6000 24GB workstation graphics card, 1TB OS SSD and 6TB SSD array, 10GbE (copper) Adapter
• EPI-FLUORESCENCE
• MICROVOLUTION (3D deconvolution method)

Laser options

• 405 nm 350mW
• 488 nm 300mW
• 560 nm 500mW
• 642 nm 500mW

Objectives lenses

• ILLUMINATION: 0.71NA water objective, 3.7mm WD
• DETECTION: 1.1NA water objective, 2.0mm WD, 62.5x total magnification

Detectors

• Dual Hamamatsu ORCA-Flash 4.0 v3 sCMOS (2k x 2k pixels with 82% QE, 100 fps)

Emission filters

• DAPI or equivalent dye 447/60 nm
• GFP, AlexaFluor488 or equivalent dye 510/42 nm or 525/50 nm
• AlexaFluor568, mCherry or equivalent dye 605/64 nm
• Cy5, AlexaFluor647 or equivalent dye 698/70 nm

Funding

This microscope was funded by an HHMI grant (PI: Dr. Joanna Wysocka). Installation date: 08/2019