Microscopy Shines a Light on Real-Time FRAP

Olympus provides the cell^frap module for the xcellence live cell imaging and manipulation system, which enables users to perform fluorescence recovery after photo-bleaching (FRAP), inverse FRAP (iFRAP), fluorescence loss in photobleaching (FLIP) and fluorescence loss after photobleaching (FLAP) experiments. The cell^frap system is also ideal for more advanced processes such as photo-conversion, photo-activation, pattern bleaching, laser cutting and trapping. The module includes its own dedicated laser system and light-path, allowing truly simultaneous imaging and photo-manipulation. These advanced techniques are made easy to control using the intuitive xcellence software interface. The cell^frap module is introduced into the microscope via a dedicated light-path and can incorporate two separate laser lines or a laser combiner. This enables simultaneous imaging and photo-manipulation, providing full integration of FRAP with epi-fluorescence, TIRFM and spinning disk confocal microscopy. The system offers unrivalled speed and precision by scanning just the region of interest, achieving a rate of bleaching up to 10 times faster than standard methods and allowing multiple discrete areas within the same frame to be bleached very quickly. This facilitates the rapid carrying-out of experiments and makes it possible to design complex protocols that investigate multiple regions of interest. In addition, intelligent hardware-based synchronisation allows μ second switching between bleaching and imaging, meaning that initial frames can be acquired as early as 150 μs after the FRAP experiment ends, providing accurate quantitative data as well as high quality images.

Fully controlled via the software, basic FRAP, FLIP and photo-switching techniques can be completed at the click of button. Additionally the ‘fire on click’ feature enables manual triggering of the FRAP laser using the mouse during time lapse acquisition. More complex analyses, such as utilising the pattern bleaching mode, can be configured easily using the intuitive interface. The refraction limited spot size of the laser enables the precise and accurate bleaching of very small structures. This set-up also allows the use of pulsed lasers to cut cellular components while simultaneously viewing the specimen, providing the capability to carry out powerful functional analyses. The Olympus xcellence systems are a new range of highly versatile, cutting-edge research microscope solutions capable of advanced live cell imaging and manipulation techniques.