Lasers for Fluorescence Microscopy

CW lasers in the visible range are widely used in fluorescence microscopy applications. The fluorescence from specific fluorophores or biomarkers is detected in confocal configurations enabling fast and high spatial resolution imaging of cells, organisms, sub-cellular structures and cell dynamics in live cells.

The resolution of these imaging techniques is fundamentally limited to ~200 nm by the diffraction limit of the laser (the Abbe principle). Over the last decades a number of so called super-resolution imaging techniques have been developed that are capable of going beyond the diffraction limit through manipulation of the fluorescence signal. Examples of super-resolution imaging techniques include STED, STORM, PALM, SIM and RESOLFT. These new techniques have enabled dynamic studies of microbiological structures at molecular level in live cells and have revolutionized the field of light microscopy.

fluorescence microscopy laser Microscopy image using GFP and RFP (image courtesy of Hjalmar Brismar. KTH Cell biology, Stockhom, Sweden)

flourence_1 Confocal microscopy principle

In a typical STED microscope, a fluorescence excitation laser beam is coaligned with a doughnut-shaped STED beam to transiently inhibit fluorescence emission by stimulated emission everywhere but at the focal center. Scanning the reduced focal spot over the sample produces images of sub-diffraction resolution. The technique enables fast imaging with a resolution that is in theory only limited by the intensity of the STED beam. A limitation of the technology has been the need for quite large and bulky high power lasers. However, in a collaboration between Cobolt and the Max Planck Institute in Göttingen, Germany, it was recently demonstrated that STED nanoscopy can also be realized with a high performance, low noise and compact CW DPSS laser. The researchers used the 500 mW output from a Cobolt Flamenco 660 nm as the STED beam and a gated detection scheme, which produced high quality live-cell STED imaging and also allowed super-resolution FCS. The demonstration shows that compact high performance DPSS lasers can strongly contribute to making the STED technology more practically and economically implementable.

Cobolt is one of the fullest range high performance laser supplier in the industry for fluorescence microscopy. The product portfolio offers a very broad selection of wavelengths across the UV/visible/near-IR spectral range, matching all typically used fluorophores, provided in compact formats and with a power scalability required for high speed imaging techniques.

See all Cobolt CW Visible 04-01 Series DPSS lasersCW UV-Visible-NIR 05-01 Series DPSS lasersultra-compact 06-01 Series lasers and Cobolt Skyra™.





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