Fully automated tcspc-flim system and time detection method based on dmd computational holographic scanning

Abstract

The invention discloses a fully automatic TCSPC-FLIM system and a time detection method based on DMD calculation holographic scanning. The continuous laser excites the sample under wide-field conditions to generate fluorescence, and the SCMOS transmits the acquired sample fluorescence information to the computer. The computer obtains the position and intensity information of the fluorescence image through calculation and processing, and selectively generates the DMD two-dimensional image corresponding to the region of interest. Hologram distribution, while controlling the DMD lens group to achieve selective light excitation. Under the control of the computer, the DMD can transmit the laser light from the first laser to the sample through the optical mirror group, the dispersion system, the DMD, etc., and excite the sample to realize FLIM. The invention applies DMD to the fully automatic TCSPC-FLIM system, adopts DMD as a scanner, and adds an automatic optical path selection system to realize fully automatic fluorescence lifetime microscopic imaging according to user definition and selection. The optical path system based on the invention greatly increases the scanning speed of imaging and realizes selective light excitation, which can effectively improve fluorescence imaging in weak fluorescence regions.

Description

technical field [0001] The present application relates to the field of biomedical imaging, in particular to a fully automatic TCSPC-FLIM system based on DMD computational holographic scanning and a time detection method. Background technique [0002] Fluorescence lifetime imaging is an extremely important direction in the field of biomedical imaging. When a substance is excited by excitation light, the molecules in the substance transition from the ground state to the first excited state after absorbing energy, and return from the excited state to the ground state. Fluorescence is emitted by means of radiative transitions. When the excitation stops, the fluorescence intensity of the fluorescent molecules will decrease, and the time taken when the intensity decreases to 1 / e of the maximum light intensity when excited is defined as the fluorescence lifetime. Two-photon is based on nonlinear optics, and its most notable features in imaging are its strong tissue penetration abi...

AI Technical Summary

Problems solved by technology

Using the fluorescence intensity imaging method is relatively straightforward, and can observe different degrees of biological imaging, but its disadvantage is that the concentration of fluorophores, the intensity of excitation light, photobleaching and the environment of fluorophores will affect the measured fluorescence intensity, which is not conducive to quantification analyze

Currently, two-photon FLIM (Fluorescence Life-time imaging Microcopy) based on TCSPC (time-correlated single photon counting) can easily visualize the fluorescence decay curve and Poisson Statistical distribution, but its imaging speed is slow and time-consuming, which is not conducive to the rapid measurement of fluorescence lifetime

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