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A three-dimensional nano-resolution positioning method and device
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A three-dimensional nanometer and positioning method technology, which is applied in material excitation analysis, fluorescence/phosphorescence, etc., can solve the problems of inability to analyze molecular positioning, low utilization rate of light energy, and high cost, and achieve the effect of improving the utilization rate of light energy.
Active Publication Date: 2015-12-02
SHENZHEN UNIV
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However, these simple three-dimensional nano-resolution positioning methods also have limitations in the imaging depth range, and cannot achieve simultaneous positioning analysis of molecules in the entire cell range.
One method of depth of field extension is to use multi-focal planes to detect the depth of field stack, using multiple detectors to image different depths in the cell, the total effective depth of field can be extended to the entire cell range, but this method has low light energy utilization efficiency , and the structure is complex, the cost is too high, etc.
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Embodiment 1
[0035] figure 1 A flow chart of the three-dimensional nano-resolution positioning method provided by the first embodiment of the present invention is shown, and for convenience of description, only parts related to this embodiment are shown.
[0036] The method mainly includes the following steps:
[0037] In step S101, perform fluorescence excitation on target molecules in the article;
[0038] In step S102, two parallel detection channels are used for fluorescence detection, and in each detection channel, the fluorescence is sequentially passed through an objective lens and a curved grating, and respectively imaged on two different detectors;
[0039] Moreover, the imaging planes of the two detection channels correspond to the dislocation distribution of the conjugate planes in the article, and the distance is 1-3 μm;
[0040] Of course, in each detection channel, a fluorescent filter can be set between the objective lens and the curved grating, a tube mirror can be set be...
Embodiment 2
[0063] Figure 4 A flow chart of the three-dimensional nano-resolution positioning method provided by the second embodiment of the present invention is shown, and for the convenience of description, only parts related to this embodiment are shown.
[0064] The method mainly includes the following steps:
[0065] In step S201, perform fluorescence excitation on target molecules in the article;
[0066] In step S202, two parallel detection channels are used for fluorescence detection, and in each detection channel, the fluorescence is sequentially passed through an objective lens and a curved grating, and imaged on different regions on the same detector;
[0067] Moreover, the imaging planes of the two detection channels correspond to the dislocation distribution of the conjugate planes in the object, and the distance is 1-3 μm;
[0068] In step S203, the image formed by the detector is analyzed by using the dual focal plane positioning method to determine the three-dimensiona...
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Abstract
The invention is suitable for the technical field of particle positioning, and provides a method for three-dimensional nanometer resolution positioning. The method comprises the following steps: conducting fluorescence excitation on target particle in an object; adopting two paralleled detection channels to conduct fluorescence detection, in each detection channel, allowing fluorescent to pass an objective lens and a curved optical grating in sequence to image in different zones of a same detector or different detectors, wherein imaging faces corresponding to the two detection channels are distributed in the staggering manner corresponding to conjugate face of the object, and the interval is 1-3 Mum; adopting dual-focal plane method to conduct analysis on images to define three-dimensional positioning information of the target particle. According to the method, the curved optical grating multi-focal plane parallel imaging and the multi-focal plane positioning method are in organic bond, so that depth of field of a conventional microscope system is broken through, and meanwhile, the three-dimensional nanometer positioning and tracking for moving particle in cell thickness area can be realized.
Description
technical field [0001] The invention belongs to the field of molecular positioning of biological cells, in particular to a three-dimensional nano-resolution positioning method and device. Background technique [0002] Cells are the basic constituent units of biological tissues and organs, and also the smallest structural and functional units of living organisms. People's understanding of complex living organisms must begin with the smallest constituent unit of life—cells. In cell biology, many dynamic life processes related to cell function are associated with the movement of vesicles and / or single molecules in cells. The tracking of biological processes in cells (such as the tracking of molecular trajectories) is an important basis for understanding cell function. For the study of these dynamic processes, one of the most used analytical methods is single-molecule tracking (SingleParticleTracking, SPT). SPT obtains nanometer-precision position information by analyzing the ...
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