Microscopic super-resolution imaging device and method based on light field multidimensional information fusion

Abstract

The invention discloses a microscopic super-resolution imaging device and method based on light field multidimensional information fusion. The device comprises a multicolor fluorescence microscopic imaging system and a single-exposure spectral imaging system, wherein the multicolor fluorescence microscopic imaging system comprises N laser devices with different wavelengths, N dichroic filters, a multichannel narrow-band filter, a three-dimensional nanometer platform used for carrying a sample, a microscope objective, a dichroic sheet, a multichannel filter and a sleeve lens; and the single-exposure spectral imaging system comprises a space random phase modulator, a relay amplifier imaging system and a photoelectric detector in sequence. Through the imaging device and method, a super-fine structure of a biological living cell can be acquired through single-exposure detection, imaging speed is high, the requirement on exciting light power density is not high, and the imaging device and method can be applied to real-time super-resolution imaging of living cells. The imaging device and method provide more significant original data for biomedical research.

Description

technical field [0001] The invention relates to fluorescence microscopic super-resolution imaging, in particular to a microscopic super-resolution imaging device and imaging method based on light field multi-dimensional information fusion. Background technique [0002] In optical microscopy imaging, the diffraction-limited resolution is determined by the numerical aperture of the microscopic objective lens and the wavelength of the incident light, and is an objective existence that cannot exceed the theoretical limit resolution. Microscopic super-resolution techniques can be divided into three categories: [0003] 1. Sparse reconstruction technology based on single-molecule localization, representative technologies include light-activated localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM) and fluorescence-activated localization microscopy (FPALM); [0004] 2. Based on point spread function engineering, it is a spatial processing technology ...

AI Technical Summary

Problems solved by technology

[0006] At present, these three types of methods have their own limitations. Wide-field imaging technologies such as STORM and PALM have a positioning accuracy of up to 20nm, but reconstructing a super-resolution image needs to collect tens of thousands of original images, and the imaging speed is limited, so it cannot be applied live cell imaging

The resolution of STED technology is determined by the ratio of the erasing optical power density to the saturation optical power density, and its imaging speed depends on the scanning speed, which is close to the imaging speed of ordinary confocal microscopy, and can be applied to live cell imaging of small fields of view. However, the STED optical power density is 4-6 orders of magnitude higher than the STORM optical power density, so the phototoxicity and photodamage to living cells cannot be ignored, and the high-power STED light also aggravates the fluorescence of fluorescent molecules while achieving fluorescence erasing. Photobleaching, which limits the application of STED in live cell imaging

Due to the limitations of the imaging principle, SIM can only double the resolution, that is, the highest is about 100nm; SSIM technology can improve the resolution to tens of nanometers comparable to STORM / PALM or STED, but the high Power density makes it lose the advantages of live cell imaging

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