Implementation And Application Of Multi-fold Pulse Splitting System

Ti:Sapphire mode-locked femtosecond laser is the key device of nonlinear imagingsystem, such as Two-photon Laser Scanning Fluorescence Microscopy. The application ofFemtosecond laser in biological imaging overcomes the strong scattering and absorptionlimitations in conventional single-photon confocal laser scanning microscopy, achieving600~800micron depth imaging in thick specimens. With the fast developing of life science,new applications demand higher imaging speed, such as fast signal detecting, long-time invivo imaging and large volume three dimension in neuron science, which couldn’t beachieved by using two-photon microscopy, due to the point excitation and high energyconcentration features. Although the way of improving laser intensity would improve thefluorescence intensity, it will increase photo-bleaching too. In the study of the femtosecondlaser itself impacting on two-photon excitation, many experiments verified that usingultra-high repetition femtosecond laser can increase the Signal to Noise Ratio(SNR), ordecrease the photo–bleaching in two-photon imaging. In this thesis, we introduce a compactmulti-fold pulse splitting device based on Passive Pulse Splitter, which can realize2-fold,4-fold,8-fold repetition increasing of femtosecond laser. Using the ultra-high repetition laserinto two-photon laser scanning fluorescence microscopy, we found that the imaging speedcan be increased eight times without SNR decreasing.Considering the output power of the laser and the energy loss of imaging system in ourlaboratory, we designed a compact multi-fold pulse splitting system, that can emissiontwo-photon fluorescence perfectly. Comparing with the two-photon imaging quality of82MHz repetition laser, we found that656MHz laser can increase the imaging speed ordecrease photon-bleaching, which is perfectly for fast signal detecting or in vivo imaging.In addition, we finished a compact multi-fold pulse splitting model that can be easilymoved and coupled into nonlinear imaging system based on femtosecond laser. This devicecan easily increase laser’s repetition outside the Femtosecond Pulse Laser, and the repetitioncan be changed quickly with using light blockers.