Microscope-based system and method for image-guided microscopic illumination
A system and method for image-guided microscopic illumination are provided. A processing module controls an imaging assembly such that a camera acquires an image or images of a sample in multiple fields of view, and the image or images are automatically transmitted to a processing module and processed by the first processing module automatically in real-time based on a predefined criterion so as to determine coordinate information of an interested region in each field of view. The processing module also controls an illuminating assembly to illuminate the interested region of the sample according to the received coordinate information regarding to the interested region, with the illumination patterns changing among the fields of view.
1 . A microscope-based system for image-guided microscopic illumination of varying patterns on a biological sample through a plurality of fields of views of the biological sample, comprising:
a microscope comprising a stage, wherein the stage is configured to be loaded with the biological sample;
an imaging assembly comprising a camera and an imaging light source;
an illuminating assembly comprising a pattern illumination device and an illuminating light source, the illuminating assembly being configured to provide an illuminating light through an illuminating light path to illuminate the biological sample;
a first processing module operatively coupled to the microscope and the imaging assembly; and
a second processing module operatively coupled to the illuminating assembly;
wherein the first processing module is configured to:
control the camera of the imaging assembly to acquire at least one image of a first field of view of the biological sample; and
process the at least one image automatically based on a predefined criterion to determine coordinate information of a region of interest in the first field of view of the biological sample;
wherein the second processing module is configured to:
receive the coordinate information of the region of interest in the first field of view of the biological sample from the first processing module;
control the illuminating assembly according to the received coordinate information to illuminate the first field of view of the biological sample with a light pattern corresponding to the region of interest in the at least one image of the first field of view of the biological sample;
wherein after the region of interest in the first field of view is illuminated, the first processing module is configured to control the stage of the microscope to move to a subsequent field of view.
2 . The microscope-based system according to claim 1 , wherein the camera is disposed on the microscope or on the imaging light path.
3 . The microscope-based system according to claim 1 , wherein the pattern illumination device comprises a pair of scanning mirrors, a digital micromirror device, or a spatial light modulator, which is disposed between the illuminating light source and the microscope along an illuminating light path.
4 . The microscope-based system according to claim 1 , wherein the first processing module comprises a computer, a workstation, or a CPU of a computer.
5 . The microscope-based system according to claim 1 , wherein the second processing module comprises a FPGA, an ASIC board, or a CPU of a computer.
6 . The microscope-based system according to claim 1 , wherein the biological sample is prepared with a photosensitizer and a chemical agent, the photosensitizer in the region of interest being excited by a illuminating light to release free radical so as to trigger the chemical agent to process a photochemical reaction.
7 . The microscope-base system according to claim 6 , wherein the photochemical reaction is biotinylation of amino acids in the region of interest.
8 . An image-guided microscopic illumination method, comprising the steps of:
a) loading a biological sample on a stage of a microscope;
b) controlling a camera and an imaging light source of an imaging assembly with a first processing module to acquire at least one image of a first field of view of a biological sample;
c) performing image processing of the at least one image automatically with the first processing module based on a predefined criterion to determine a region of interest in the at least one image and obtain coordinate information of the region of interest;
d) transmitting the coordinate information of the region of interest in the at least one image to a second processing module;
e) controlling an illuminating light source and a pattern illumination device of an illuminating assembly with the second processing module according to the coordinate information to illuminate the first field of view of the biological sample with a light pattern corresponding to the region of interest in the at least one image of the first field of view of the biological sample; and
f) after the region of interest in the first field of view is illuminated, controlling the stage of the microscope with the first processing module to move to a subsequent field of view.
9 . The image-guided microscopic illumination method according to claim 8 , wherein after controlling the stage to the subsequence of view, repeating the steps b) through f), until regions of interest in all subsequent fields of view are illuminated.
10 . The image-guided microscopic illumination method according to claim 8 , wherein the image processing is performed with real-time image processing techniques comprising one or more of thresholding, erosion, filtering, or artificial intelligence trained semantic segmentation methods.