Total internal reflection illumination and semi-total internal reflection illumination dual optical path fluorescence microscope system

Abstract

The invention discloses a total internal reflection illumination and semi-total internal reflection illumination dual optical path fluorescence microscope system. A beam-splitter plate I is arranged on the light path of exciting light and the angle between the beam-splitter plate I and the light path is 135 degrees so that the exciting light is divided into a vertical transmission light and a vertical reflected light; the transmission light is reflected through a reflector so as to be parallel to the reflected light; the light paths of the reflected light and the transmission light are respectively and sequentially provided with a graded-density neutral filter, a convex lens I, a convex lens II and a light spot aperture regulator; the reflected light and the transmission light then form two vertical light paths by adjusting transmission directions through the reflector; convex lens III are respectively arranged on the mutually vertical light paths of the reflected light and the transmission light; the reflected light and the transmission light pass through the convex lens III so as to be incident to a beam-splitter plate II having an angle of 135 degrees with a light path of the reflected light; and two parallel lights divided through the beam-splitter plate II are incident to an exciting light inlet of a microscope. The total internal reflection illumination and semi-total internal reflection illumination dual optical path fluorescence microscope system can be used for bleaching fluorescence molecule of a cell membrane area, background fluorescence interference caused when the fluorescence molecule of the cell membrane area images a focal plane in a cell is avoided, and a signal background ratio of semi-total internal reflection fluorescence imaging in the cell is further improved.

Description

technical field [0001] The invention relates to a fluorescent microscopic system with two optical paths of total internal reflection illumination and semi-total internal reflection illumination, which belongs to the field of fluorescence microscopic imaging. Background technique [0002] Life science is gradually moving from macro to micro. Realizing the imaging and characterization of proteins, nucleic acids and other biomolecules under physiological conditions at the subcellular level is of great significance for revealing the mysteries of life and improving the prevention, diagnosis and treatment of diseases. [0003] Fluorescence imaging is the most commonly used imaging characterization technique. In cell imaging, since fluorescent labels are often not limited to the focal plane, when imaging with a common wide-field fluorescence microscope, illuminants outside the focal plane often bring strong background fluorescence. interference, which limits its application in subc...

AI Technical Summary

Problems solved by technology

Since the fluorescence in the cell membrane area will also interfere with the imaging of the intracellular area, the signal-to-background ratio is not significantly improved, and the application of semi-total internal reflection fluorescence microscopy is limited to a certain extent.

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