CN105353514B - Reduce method, the method and apparatus of laser microprobe dating of laser beam cross-section product - Google Patents

Abstract

The invention provides a method for reducing the cross-sectional area of the laser beam. A spatial light modulator is arranged in the optical path between the laser emitter and the object to be illuminated; the incident laser beam emitted by the laser emitter The main light spot and the auxiliary light spot are generated; the main light spot covers N adjacent micro-units of the spatial light modulator, and only n micro-units centered on the center of the main light spot are opened (n=1,2,...,N ), and the other N-n micro-units are in the off state, then the cross-sectional area of the outgoing laser beam d=D*(n/N), where D is the cross-sectional area of the main spot. The invention also provides a laser microdissection method and device. The above-mentioned method for reducing the cross-sectional area of the laser beam reduces the cross-sectional area of the laser beam within a certain range through manual operation and control. The above-mentioned laser microdissection method and device can realize laser cutting with a cutting line width as small as micron or even submicron.

Description

Method for reducing cross-sectional area of laser beam, method and device for laser microdissection

technical field

The invention relates to the field of laser cutting, in particular to laser micro-cutting.

Background technique

In life science research, it is necessary to conduct gene and protein research on a specific cell (group) in a tissue. The laser micro-cell dissection system is a high-end equipment developed to meet such needs. It can separate a specific or characteristic cell (group) from a variety of tissues, thereby preventing the experimental specimen from being Contamination by other cells, bacteria, or other impurities makes the analysis of genes and proteins more accurate and specific.

In the laser microscopic cell dissection technology, it is sometimes necessary to cut a single cell: first, observe and judge under a microscope to determine the cells that need to be selected; secondly, determine the cell cutting line according to the cell shape; Simultaneously, the stage is driven to move the glass slide with the cell sample to generate a closed cutting line track. In this way, the laser produces a closed cutting trajectory along the cutting line around the cell, so that the cell is separated from other cells in the surrounding background. Afterwards, the cells can be separated by collection systems such as gravity or electrostatic adsorption, so as to obtain pure cells without impurities.

The size of the cell is between a few microns to more than 10 microns, which is very small. Sometimes it is necessary to cut and separate a segment of gene selected by the user (included in the cell), and the size of the gene is even smaller. Therefore, a "knife" for cutting cells is needed: the cross-sectional area of the laser beam is very small, on the order of submicron or micron, especially when it is necessary to separate and cut the gene fragments contained in the cells selected by the user. The cross-section of a general laser beam spot is approximately a circle or an ellipse, and the scale along one direction is on the order of 100 microns or even millimeters. Therefore, how to realize the laser beam whose cross-section length is micron or even submicron is a difficult problem.

Contents of the invention

The main technical problem to be solved by the present invention is to provide a method for reducing the cross-sectional area of the laser beam. Through manual operation and control, the cross-sectional area of the laser beam can be reduced within a certain range.

Another main technical problem to be solved by the present invention is to provide a method and device for laser microdissection, which can realize laser cutting with micron or even submicron diameter.

In order to solve the above-mentioned technical problems, the present invention provides a method for reducing the cross-sectional area of the laser beam. A spatial light modulator is arranged in the optical path between the laser emitter and the object to be illuminated; the incident light emitted by the laser emitter The laser beam produces a main spot and a side spot on the spatial light modulator;

The main light spot covers N adjacent micro-units of the spatial light modulator, only n micro-units (n=1, 2,...,N) centered on the center of the main light spot are turned on, and the other N-n micro-units are turned off State, then the cross-sectional area of the outgoing laser beam d=D*(n/N), where D is the cross-sectional area of the main spot.

In a preferred embodiment: the micro-unit of the spatial light modulator covered by the auxiliary light spot is in an off state.

The present invention also provides a method for laser microdissection, in which a spatial light modulator is arranged in the optical path between the laser emitter and the objective lens; the incident laser beam emitted by the laser emitter produces a main light spot and a secondary spot;

The main light spot covers N adjacent micro-units of the spatial light modulator, only n micro-units (n=1, 2,...,N) centered on the center of the main light spot are turned on, and the other N-n micro-units are turned off State, then the cross-sectional area of the outgoing laser beam after passing through the objective lens d=D*n/(N*M), where D is the cross-sectional area of the main spot, and M is the magnification of the objective lens.

In a preferred embodiment: the micro-unit of the spatial light modulator covered by the auxiliary light spot is in an off state.

The present invention also provides a laser microdissection device, including a microscope, a microscope illumination system, a digital imaging system, and a laser emitter; A spatial light modulator; the incident laser beam emitted by the laser emitter produces a main spot and a secondary spot on the spatial light modulator;

The main light spot covers N adjacent micro-units of the spatial light modulator, only n micro-units (n=1, 2,...,N) centered on the center of the main light spot are turned on, and the other N-n micro-units are turned off State, then the cross-sectional area of the outgoing laser beam after passing through the objective lens d=D*n/(N*M), where D is the cross-sectional area of the main spot, and M is the magnification of the objective lens.

In a preferred embodiment: the micro-unit of the spatial light modulator covered by the auxiliary light spot is in an off state.

In a preferred embodiment: the laser beam optical path shaping system further includes a dichroic mirror, which reflects the cutting laser to the objective lens, and transmits the imaging objective lens to the digital imaging system.

In a preferred embodiment: the microscope is an upright microscope or an inverted microscope

Description of drawings

Fig. 1 is the optical path diagram of preferred embodiment 1 of the present invention;

Fig. 2 is the optical path schematic diagram of preferred embodiment 2 of the present invention;

Fig. 3 is the optical path schematic diagram of preferred embodiment 3 of the present invention;

Fig. 4 is a schematic diagram of the optical path of the preferred embodiment 4 of the present invention.

Detailed ways

The method of the present invention to realize the laser beam whose cross-sectional area is submicron or micron in length is to use a spatial light modulator. There are mainly two types of spatial light modulators: transmissive and reflective, and their main feature is the "optical switch" with spatial distribution. The main function is to selectively reflect (or transmit) the optical signals irradiated at some positions of the spatial light modulator, while the optical signals at other positions cannot be reflected (or transmitted).

Introduction to using DMD as a spatial light modulator: DMD was invented by Dr. Larry Hornbeck of TI in 1987. It uses MEMS (micro-electromechanical systems) technology to integrate micromirror arrays and CMOS SRAM on the same chip. A new, all-digital spatial light modulator. DMD is composed of millions of micro-units. A DMD with 1024X768 micro-units has a diagonal length of only 0.7 inches and a thickness of about 5 mm (as shown in Figure 1). Each microunit unit is mainly composed of microunits, hinges and CMOS substrates, in which each microunit is connected by a yoke plate and a torsional hinge, which can rotate between two angles of -12° and +12° with the hinge as the axis , and its rotation direction is determined by the voltage of the controllable electrodes on both sides of the yoke plate. The electrode of each microcell has a storage unit SRAM, which is located under the platform of the controllable electrode. When the SRAM stores "1", the microcell deflects +12 °, when "0" is stored, the deflection is -12°. When designing the optical system of DMD, generally the light of a certain reflection direction of the micro-unit enters the optical system, which means "open" (assuming it is +12° direction, the following is similar), and the light of another reflection direction will not enter the optical system System, means "off". Therefore, DMD can be simply regarded as an optical switch array, and "1" or "0" is stored in the corresponding SRAM unit according to the need, and the switch of the corresponding micro unit can be controlled to generate any pattern.

It should be noted that although the DMD is used as an example above, the solution of the present invention is not limited to the DMD as a spatial light modulator. The present invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

Example 1

A method for reducing the cross-sectional area of a laser beam. A spatial light modulator is arranged in the optical path between a laser emitter and an object to be illuminated; the incident laser beam emitted by the laser emitter generates a main spot on the spatial light modulator and secondary facula;

Referring to Fig. 1, the main light spot covers N adjacent micro-units of the spatial light modulator, and only n micro-units (n=1, 2, ..., N) centered on the center of the main light spot are opened, and the other N-n If a micro unit is in the off state, then the cross-sectional area of the outgoing laser beam d=D*(n/N), where D is the cross-sectional area of the main spot. Therefore, the cross-sectional area of the outgoing laser beam can be reduced, and the cross-sectional area of the outgoing laser beam can be changed within a certain range by changing the number of closed micro-units. Therefore, the purpose of simply changing the cross-sectional area of the laser beam is achieved.

Furthermore, the micro-units of the spatial light modulator covered by the secondary light spots are in the off state. In this way, the emitted laser light of the sub-spot is completely separated from the emitted laser light of the main spot, so that the sub-spot is eliminated.

Example 2

With reference to Fig. 2, a kind of device of laser microdissection comprises upright microscope, microscope illumination system, digital imaging system and laser emitter 1; The spatial light modulator 3 in the optical path of the objective lens 2; the incident laser beam emitted by the laser emitter 1 produces a main spot and a secondary spot on the spatial light modulator 3;

The main light spot covers N adjacent micro-units of the spatial light modulator, and only n micro-units (n=1, 2, ..., N) centered on the center of the main light spot are opened, and the other N-n micro-units are In the closed state, the cross-sectional area of the outgoing laser beam passing through the objective lens d=D*n/(N*M), where D is the cross-sectional area of the main spot, and M is the magnification of the objective lens.

Assume that the cross-sectional area of the main spot of the laser beam generated by the laser emitter 1 is D=100um*100um, and the magnification of the objective lens 2 is M=100 times. The side length of the micro-unit is about 10 microns. Therefore, the number of micro-units covered by the main light spot is N=10*10. Then the above-mentioned laser microdissection device realizes the cross-sectional area of the minimum outgoing laser beam d=0.1 micron*0.1 micron, that is, only one micro-unit at the center of the main spot is opened, so that n=1. That is, the width of the cutting line reaches 0.1 micron. Considering the diffraction effect, the size of the light spot on the cell sample is also at the sub-micron level, which can meet the needs of single-cell or even sub-cellular cutting.

Furthermore, the micro-units of the spatial light modulator 3 covered by the auxiliary light spot are in the off state. In this way, the emitted laser light of the sub-spot is completely separated from the emitted laser light of the main spot, so that the sub-spot is eliminated.

Furthermore, the laser beam optical path shaping system also includes a dichroic mirror 5, which reflects the cutting laser to the objective lens and transmits the imaging objective lens to the digital imaging system.

In this embodiment, the optical system of the illumination source includes a condenser 6 and an illumination source 7; the light emitted by the illumination source 7 passes through the condenser 6 and then irradiates to the motorized stage 8, and the side of the motorized stage 8 away from the illumination source is placed with Cut target 9. The outgoing laser beam emitted by the objective lens 2 is directly irradiated on the cutting target 9 .

Example 3

The difference between this embodiment and embodiment 2 is that the upright microscope is replaced with an inverted microscope. Referring to FIG. 3 , it is only necessary to place the cutting target 9 on the side of the motorized stage 8 facing the epi-illumination light source 7 . The outgoing laser beam emitted by the objective lens 2 passes through the motorized stage 8 and irradiates on the cutting target 9 .

Example 4

The difference between this embodiment and Embodiment 3 is that the epi-illumination light source is replaced by the transmission illumination light source 7. Referring to FIG. A cutting target 9 is placed on one side of the transmission illumination light source 7 . The outgoing laser beam emitted by the objective lens 2 passes through the motorized stage 8 and irradiates on the cutting target 9 .

The above is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed in the present invention can easily think of changes or Replacement should be covered within the protection scope of the present invention. Therefore, the protection scope of the present invention should be determined by the protection scope of the claims.

Claims (5)

1. a kind of method reducing laser beam cross-section product, it is characterised in that：Light between laser emitter and illuminated object Installation space optical modulator in road；The incoming laser beam that the laser emitter is sent out generates main spot in spatial light modulator With secondary hot spot；

The main spot covers on N number of adjacent micro unit of spatial light modulator, only opens the n centered on the main spot center of circle Micro unit, n=1,2 ..., N, other N-n micro units are in off state, and the micro unit covered by secondary hot spot is to close shape State, then the cross-sectional area d=D* (n/N) of shoot laser beam, wherein D are the cross-sectional area of main spot.

2. a kind of method of laser microprobe dating, it is characterised in that：The installation space light in the light path of laser emitter and object lens Modulator；The incoming laser beam that the laser emitter is sent out generates main spot and secondary hot spot in spatial light modulator；

The main spot covers on N number of adjacent micro unit of spatial light modulator, only opens the n centered on the main spot center of circle Micro unit, n=1,2 ..., N, other N-n micro units are in off state, and the micro unit covered by secondary hot spot is to close shape State.

3. a kind of device of laser microprobe dating, including microscope, microscope illumination system, digital imaging system and Laser emission Device, it is characterised in that further include laser beam optical path orthopedic systems comprising one is set to laser emitter and micro objective light Spatial light modulator in road；The incoming laser beam that the laser emitter is sent out generated in spatial light modulator main spot and Secondary hot spot；

The main spot covers on N number of adjacent micro unit of spatial light modulator, only opens the n centered on the main spot center of circle Micro unit, n=1,2 ..., N, other N-n micro units are in off state, and the micro unit covered by secondary hot spot is to close shape State.

4. a kind of device of laser microprobe dating according to claim 3, it is characterised in that：The laser beam optical path shaping System further includes a dichroscope, will cutting laser reflection to object lens, number will be transmitted through by the image optics signal of object lens Imaging system.

5. a kind of device of laser microprobe dating according to any one of claim 3-4, it is characterised in that：It is described micro- Mirror is just to set microscope or inverted microscope.