Method and apparatus for carrying out wide-field three-dimensional ultra-high resolution positioning and imaging on intact cell

Abstract

The invention discloses a method for carrying out wide-field three-dimensional ultra-high resolution positioning and imaging on an intact cell. The method comprises: modulating the polarization state of incident light into P-polarized light or S-polarized light; illuminating a sample after the modulated light beam becomes into tangential polarized light or radial polarized light; illuminating the lower surface of the cell by using the evanescent wave generated by light having an incident angle of more than a total internal reflection critical angle; illuminating the upper surface of the cell by using a pattern, wherein transmitted light having an incident angle of less than a total internal reflection critical angle and the reflected light generated after one reflection with the upper surface of the cell are subjected to interference to generate the pattern; and according to the fluorescence information emitted by the collected sample, re-constructing the three-dimensional super-resolution image of the intact cell. The invention further discloses an apparatus for carrying out wide-field three-dimensional ultra-high resolution positioning and imaging on an intact cell. According to the present invention, the method and the apparatus have characteristics of rapid imaging, simple equipment, convenient operation and the like, and can be well used for the detection of fluorescent samples.

Description

technical field [0001] The invention belongs to the field of super-resolution microscopic imaging, in particular to a wide-field three-dimensional super-high-resolution positioning and imaging method and device for intact cells. Background technique [0002] Conventional wide-field optical microscopes are affected by diffraction effects, and the lateral resolution is limited to about half the wavelength of the excitation light, and the axial resolution is even lower. To understand the basis of molecular regulation in some complex biological processes, analysis of the dynamics of proteins in living cells in cultured cells or intact tissues is required. [0003] The key to achieve lateral super-resolution in many commonly used imaging methods at present is to modulate the intensity of the fluorescent signal, such as the structural intensity modulation in STED and SSIM, and the intensity modulation in the random process in PALM and STORM. These intensity modulations are depend...

AI Technical Summary

Problems solved by technology

However, the imaging range of TIRF is limited to a few hundred nanometers near the cover glass; at the same time, the acquisition of axial information by TIRF is still a positioning technology, and cannot obtain three-dimensional super-high resolution information.

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