Fluorescence microscopy is a balance between light dose, resolution, signal-to-noise, and field of view. Total Internal Reflection Fluorescence (TIRF) microscopy uses an evanescent wave of energy immediately adjacent to the coverslip to excite fluorophores in the specimen. The excitation occurs about 100nm from the coverslip, which is perfect for live cell imaging including focal adhesions, membrane dynamics, receptor function and single-molecule in vitro imaging. TIRF requires placing a narrowly focused beam at the edge of the back focal plane of an objective with NA greater than 1.38 and typically has some shadowing and direction-related artifact in the resulting evanescent wave illumination. Spinning the beam via galvo mirrors around the periphery of the back focal plane at high-speed averages out any artifacts and results in smooth, evenly illuminated images.
TIRF microscopy creates an evanescent wave that is about 100nm thick above the coverslip leading to a very thin plane of excitation, even when compared to lightsheet microscopy. This allows scientists to image events on the glass surface with essentially no background when compared to traditional widefield imaging.
VectorTIRF allows for TIRF imaging with high speed and unrivaled clarity on a comparatively large, evenly illuminated field via spinning the excitation light around the back focal plane. Spinning of the excitation light leads to more uniform imaging of thin samples without shadowing and polarization artifacts, as seen in the diagram on the right. VectorTIRF can be used for a variety of live-cell, in vitro, and even single molecule imaging when combined with a modern sCMOS or EMCCD detector. The flexibility of VectorTIRF allows integration with all research inverted microscope frames.
Standard VectorTIRF (left) and thunderSTORM reconstruction (right) of Alexa647-labeled microtubles in COS7 cells.
• Up to 200mW lasers
• Fiji direct import of SlideBook .sld files
405nm, 445nm, 488nm, 515nm, 561nm, 594nm, 640nm
SlideBook software for image acquisition and analysis
Polarization input adapter for imaging with two orthogonal polarized input beams
10 mm with variable beam expansion for fine tuning of excitation power
Mounts to the sideport on Zeiss, Nikon, Olympus, and Leica inverted microscopes