Dynamic super-resolution fluorescence imaging technology with adaptive sampling frame rate

Abstract

The invention provides a dynamic super-resolution fluorescence imaging technology with adaptive sampling frame rate. The invention relates to a dynamic super-resolution fluorescence microscopic imagecapturing technology. (1) A fluorescence microscopic imaging device is capable of super-resolution imaging and low-resolution imaging, and the field of view regions and object planes observed by two imaging modes are the same. The device can process a low-resolution image in real time and carries out feedback control on the sampling frame rate of a super-resolution image. (2) A dynamic super-resolution fluorescence microscopic image capturing method corresponding to the device comprises the steps that the low-resolution image is collected at a fixed high-speed sampling frame rate for real-timeprocessing; the dynamic change rate of the image is analyzed by calculating the optical flow; the dynamic change rate of the image is converted into the acquisition time interval of the super-resolution image; and the sampling frame rate of the super-resolution image is controlled in real time through a feedback loop, so that the super-resolution image can adapt to the dynamic change rate of a sample. The device and the method are used for realizing super-resolution capturing of a rapid dynamic change process of the sample, and can prolong the capturing time of the sample as much as possible.

Description

technical field [0001] The invention relates to the technical field of super-resolution fluorescence imaging, in particular to a shooting technology of dynamic super-resolution fluorescence microscopic images. Background technique [0002] Traditional fluorescence microscopes (such as wide-field illumination, scanning confocal, total internal reflection illumination, etc.) are limited by the Abbe diffraction limit, and their spatial resolution cannot be better than 200nm, so they cannot distinguish finer structures. In the past ten years, a variety of fluorescence imaging methods that break through the diffraction limit have been developed, called super-resolution imaging, such as stimulated emission depletion imaging (STED), reversible saturation / switching optical transition imaging (RESOLFT), and photoactivated localization microscopy. Imaging (PALM / FPALM), Stochastic Optical Reconstruction Microimaging (STORM), Structured Illumination Microimaging (SIM, Saturated Structur...

AI Technical Summary

Problems solved by technology

[0002] Traditional fluorescence microscopes (such as wide-field illumination, scanning confocal, total internal reflection illumination, etc.) are limited by the Abbe diffraction limit, and their spatial resolution cannot be better than 200nm, so they cannot distinguish finer structures

In the past ten years, a variety of fluorescence imaging methods that break through the diffraction limit have been developed, called super-resolution imaging, such as stimulated emission depletion imaging (STED), reversible saturation / switching optical transition imaging (RESOLFT), and photoactivated localization microscopy. Imaging (PALM / FPALM), Stochastic Optical Reconstruction Microimaging (STORM), Structured Illumination Microimaging (SIM, Saturated Structured Illumination SSIM, Nonlinear Structured Illumination NSIM), etc.; these super-resolution imaging techniques require more Higher intensity laser illumination (such as STED, SSIM, NSIM, etc.), or the need to repeatedly turn on / off fluorescent molecules (such as RESOLFT, PALM / FPALM, STORM, etc.), severely aggravates the fluorescence bleaching and phototoxicity of the sample; therefore, When these imaging methods with spatial super-resolution capabilities are applied to the imaging of dynamic processes in living cells, the number of image frames that can be continuously captured is far less than that of traditional fluorescence imaging methods

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