CN102540439B - Confocal axial scanning device and confocal axial scanning method based on reflection type liquid crystal spatial light modulator - Google Patents

Abstract

A confocal axial scanning device and a confocal axial scanning method based on a reflective liquid crystal spatial light modulator relate to a confocal microscope and a scanning method thereof. Defocusing is achieved by moving the objective lens or the object to be measured in the axial direction, resulting in a complex structure of the device and poor repeatability of the measurement of the sample. The confocal axial scanning device of the present invention adds a reflective liquid crystal spatial light modulator between the polarization beam splitter and the objective lens on the basis of the existing device, and removes the axial scanning mechanism in the traditional confocal axial scanning device at the same time. A liquid crystal spatial light modulator replaces the axial scanning mechanism. The confocal axial scanning method of the present invention adopts the method of directly modulating the phase grayscale image by the computer to adjust the wavefront of the light wave before the objective lens, so as to realize zooming, so that neither the measuring objective lens nor the measured object can move axially to realize the measurement of the measured object. Axial scanning of the measured object. For confocal microscopy and its scanning.

Description

Confocal axial scanning device and confocal axial scanning method based on reflective liquid crystal spatial light modulator

technical field

The invention relates to a confocal microscope and a scanning method thereof.

Background technique

Since the 1980s, microelectronics technology, bioengineering, micro-optics and micro-opto-electromechanical system technology have entered a stage of rapid development. In the field of micro-scale, people pay more attention to the quantitative analysis of three-dimensional state. Ultra-precise measurement has become an important topic in modern testing technology and instrument research. The confocal measurement method has become an important research direction of three-dimensional microstructure measurement because of its unique advantages of high precision, high resolution, non-contact and easy realization of three-dimensional imaging digitization. One method of using confocal measurement method to realize three-dimensional microstructure measurement is to make the measured object move axially relative to the focus position, form a defocus signal on the surface of the detector, and realize the measurement of the measured object according to the light intensity detected by the detector. Measurement of the three-dimensional microstructure of objects. A typical measurement setup is as follows:

Dr. Wang Fusheng from the Department of Automation Test and Control of Harbin Institute of Technology proposed a confocal device that can realize axial scanning in his doctoral thesis "Research on Three-dimensional Measurement System Based on Differential Confocal Microscopic Detection Technology". The device is composed of Including: laser, converging lens, pinhole 3, beam expander, polarizing beam splitter, λ/4 wave plate, objective lens, object to be measured, also includes: condenser, beam splitter, pinhole 1, photodetector 1, pinhole 2. Photodetector 2. The parallel light beam emitted by the laser is imaged by the converging lens to the pinhole 3 to form a point light source. The light beam emitted by the point light source is parallel incident on the polarization beam splitter through the beam expander, and the light beam transmitted through the polarization beam splitter passes through the λ/4 wave plate The image is imaged by the objective lens to the surface of the measured object and reflected along the original path. After passing through the λ/4 wave plate again, it is reflected by the polarizing beam splitter to the condenser to form a converged beam. The two pinholes behind the focus and focus are detected by a photodetector to form a differential confocal system. The device realizes axial scanning by moving the objective lens forward and backward along the optical axis direction through the Z-direction drive unit.

Dr. Huang Xiangdong from the Department of Automation Test and Control of Harbin Institute of Technology introduced the working principle of the reflective laser scanning confocal microscope in his doctoral thesis "Array Spatial Super-resolution Confocal Microscanning Detection Technology and Theoretical Research". The device consists of : laser, pinhole 1, beam splitter, objective lens, object to be measured, also includes: pinhole 2, detector. The beam emitted by the laser passes through the pinhole 1 to form a point light source. The beam emitted by the point light source is imaged by the objective lens to the surface of the measured object and reflected along the original path. The reflected light passes through the objective lens again and is reflected by the beam splitter to the position of the pinhole 3. The device realizes the system's axial scanning of the measured object by adjusting the axial position of the measured object.

These two doctoral thesis both proposed a confocal microscopy technique and scanning method that can realize axial scanning. Their common feature is that the defocusing of the measured object is realized by moving the objective lens or the measured object along the axial direction. The method requires the measurement device to include an axial scanning mechanism. The first disadvantage of the device with an axial scanning mechanism is that the structure of the device is complex, and the movement mechanism, control mechanism, and feedback link are required to achieve precise measurement; the second disadvantage is that if the sample to be measured is moved axially, and the sample is Deformation will continue to occur during the movement, such as cells, which not only reduces the measurement accuracy of such samples, but also poor measurement repeatability.

Contents of the invention

The purpose of the present invention is to solve the existing axial scanning device and method because the defocusing of the measured object is realized by moving the objective lens or the measured object along the axial direction, resulting in complex structure of the device and poor measurement repeatability of the sample. The problem is to provide a confocal axial scanning device and a confocal axial scanning method based on a reflective liquid crystal spatial light modulator.

A confocal axial scanning device based on a reflective liquid crystal spatial light modulator, which includes a laser, a converging lens, a first pinhole, an illumination objective lens, a diaphragm, a polarizing beam splitter, a λ/4 wave plate, and a reflective liquid crystal spatial light modulation The parallel light beam emitted by the laser is imaged to the first pinhole through the converging lens, and the light beam passing through the first pinhole is collimated by the illumination objective lens. The polarizing beam splitter divides the incident light into reflected light and transmitted light, the reflected light is S light, the transmitted light is P light, and the transmitted light reaches the reflection after passing through the λ/4 wave plate The reflective liquid crystal spatial light modulator totally reflects the arriving light, and the obtained total reflected light returns to the polarizing beam splitter according to the original optical path, and the reflected light reflected by the polarizing beam splitting mirror enters the beam splitting mirror, and then passes through the polarizing beam splitter. The reflected light reflected by the beam splitter enters the measuring objective lens, and is imaged on the measured object through the measuring objective lens. The reflected light reflected by the measured object returns to the beam splitter along the original optical path, and the transmitted light transmitted by the The objective lens images the image at the position of the second pinhole, and enters the photosensitive surface of the light intensity detector through the second pinhole.

Based on the above-mentioned confocal axial scanning device based on the reflective liquid crystal spatial light modulator to realize the confocal axial scanning method, its specific steps are as follows:

Step 1. Set the theoretical focusing distance x _F = 0, configure the object to be scanned as the object to be scanned, and place the object to be scanned at a distance of 2f ₀ from the measurement objective lens;

Step 2. Keep the relative position of the object to be scanned and the measuring objective lens described in step 1 unchanged, and control the change of the phase grayscale image of the reflective liquid crystal spatial light modulator through the computer, so that the theoretical focusing distance corresponding to the phase grayscale image is x _F along the axial direction from 0 μm to 105 μm in steps of 5 μm, the relationship between the phase grayscale image and the theoretical focusing distance x _F is:

Wherein, _f0 is the focal length of the measuring objective lens, d is the optical path from the reflective liquid crystal spatial light modulator to the measuring objective lens, λ is the wavelength of the light wave, and ξ and η represent two-dimensional space coordinates;

After adjusting the phase grayscale image of the reflective liquid crystal spatial light modulator each time, use the light intensity detector to image to obtain the corresponding light intensity information;

Step 3: Obtain a corresponding confocal axial scanning curve according to each theoretical focusing distance x _F and light intensity information obtained in step 2.

The confocal axial scanning device of the present invention adds a reflective liquid crystal spatial light modulator between the polarization beam splitter and the objective lens on the basis of the existing device, and simultaneously removes the axial scanning mechanism in the traditional confocal axial scanning device , replace the axial scanning mechanism with a reflective liquid crystal spatial light modulator. The structure of the device of the present invention is simpler than that of the prior art. The confocal axial scanning method of the present invention uses a computer to directly modulate the phase grayscale image to adjust the wavefront of the light wave in front of the objective lens, thereby realizing zooming, so that the measurement objective lens and the object are measured. The measured object can realize the axial scanning of the measured object without axial movement, which not only avoids the precise and complicated control of the three-dimensional micro-displacement platform, but also makes the confocal axial scanning method described in the present invention easier to operate. At the same time, the measurement accuracy and repeatability of deformable samples are improved, thereby expanding the types of sample measurements.

Description of drawings

Fig. 1 is a schematic structural diagram of a confocal axial scanning device based on a reflective liquid crystal spatial light modulator, Fig. 2 is a phase grayscale diagram of a reflective liquid crystal spatial light modulator when the focusing distance x _F = 10 μm, and Fig. 3 is a focus adjustment The phase grayscale image of the reflective liquid crystal spatial light modulator when the distance x _F =50 μm, Figure 4 is a schematic diagram of the confocal axial response experimental device based on the reflective liquid crystal spatial light modulator, Figure 5 is the focus distance of 40 μm and Comparison of confocal axial scanning curves without focusing, in which the solid line is the curve after the focusing distance is 40 μm, and the dotted line is the curve without focusing.

Detailed ways

Specific Embodiment 1: This embodiment is described in conjunction with FIG. 1. The confocal axial scanning device based on a reflective liquid crystal spatial light modulator described in this embodiment includes a laser 1, a converging lens 2, a first pinhole 3, and an illumination objective lens. 4. Aperture 5, polarizing beam splitter 6, λ/4 wave plate 7, reflective liquid crystal spatial light modulator 8, measured object 10, measuring objective lens 11, beam splitter 12, collecting objective lens 13, second pinhole 14 and The light intensity detector 16, the parallel light beam emitted by the laser 1 is imaged to the first pinhole 3 through the converging lens 2, the light beam passing through the first pinhole 3 is collimated by the illumination objective lens 4, and then enters the polarization beam splitter 6 in parallel, and the polarization beam splitter The mirror 6 divides the incident light into reflected light and transmitted light, the reflected light is S light, the transmitted light is P light, and the transmitted light reaches the reflective liquid crystal spatial light modulator 8 after passing through the λ/4 wave plate 7, and the reflected light The liquid crystal spatial light modulator 8 totally reflects the arriving light, and the total reflected light obtained returns to the polarization beam splitter 6 according to the original optical path, and the reflected light reflected by the polarization beam splitter 6 enters the beam splitter 12, and passes through the beam splitter 12 The reflected reflected light is incident on the measuring objective lens 11, and is imaged on the measured object 10 through the measuring objective lens 11. The light is imaged by the collecting objective lens 13 to the position of the second pinhole 14 , and enters the photosensitive surface of the light intensity detector 16 through the second pinhole 14 .

Specific Embodiment 2: This embodiment is described in conjunction with FIG. 1. The difference between this embodiment and the confocal axial scanning device based on a reflective liquid crystal spatial light modulator described in Embodiment 1 is that it also includes a displacement stage 9, and the measured The object 10 is placed on the translation platform 9, and the translation platform 9 can move one-dimensionally along the optical axis direction of the measuring objective lens 11. Other components and connections are the same as those in Embodiment 1.

Specific Embodiment 3: This embodiment is described in conjunction with FIG. 1. The difference between this embodiment and the confocal axial scanning device based on a reflective liquid crystal spatial light modulator described in Embodiment 1 is that it also includes a three-dimensional precision translation stage 15, The light intensity detector 16 is placed on the three-dimensional precision translation platform 15, and the three-dimensional precision translation platform 15 can move in three dimensions. Other components and connections are the same as those in Embodiment 1.

Embodiment 4: The effective pixel area of the reflective liquid crystal spatial light modulator 8 described in Embodiment 1 is 7.68 mm×7.68 mm, and the size of each pixel is 15 μm×15 μm.

Embodiment 5: Using the confocal axial scanning device based on the reflective liquid crystal spatial light modulator described in Embodiment 1 to realize the confocal axial scanning method, it includes the following specific steps:

Step 1. Set the theoretical focusing distance x _F = 0, configure the object to be scanned as the object to be scanned, and place the object to be scanned at a position where the distance from the measuring objective lens 11 is 2f ₀ ;

Step 2. Keep the relative position of the object to be scanned and the measuring objective lens 11 described in step 1 unchanged, and control the change of the phase grayscale map of the reflective liquid crystal spatial light modulator 8 through the computer, so that the theoretical adjustment corresponding to the phase grayscale map The focal distance x _F is from 0 μm to 105 μm in steps of 5 μm along the axial direction, and the relationship between the phase grayscale image and the theoretical focusing distance x _F is:

Wherein, _f0 is the focal length of the measuring objective lens, d is the optical path from the reflective liquid crystal spatial light modulator to the measuring objective lens, λ is the wavelength of the light wave, and ξ and η represent two-dimensional space coordinates;

After adjusting the phase grayscale image of the reflective liquid crystal spatial light modulator 8 each time, the light intensity detector 16 is used for imaging to obtain the corresponding light intensity information;

Step 3: Obtain a corresponding confocal axial scanning curve according to each theoretical focusing distance x _F and light intensity information obtained in step 2.

Embodiment 6: In conjunction with Fig. 4, this embodiment is a specific embodiment of Embodiment 5. In this embodiment, a stepper motor is used in a confocal axial scanning device based on a reflective liquid crystal spatial light modulator. The drive of displacement table 9, described stepping motor adopts SC300-2A type stepping motor driver to drive;

The method is:

Step 1. Set the theoretical focusing distance x _F =0, and adjust the image sensor to the focal position of the measuring objective lens 11;

Step 2. According to the selected theoretical focusing distance x _F of 5 μm, 10 μm, 15 μm, 20 μm, 25 μm, 30 μm, 35 μm, 40 μm, 45 μm, 50 μm, 75 μm, 100 μm, 175 μm and 250 μm, control the reflective liquid crystal space through the computer The light modulator 8, so that the phase grayscale image of the reflective liquid crystal spatial light modulator 8 satisfies:

Wherein, _f0 is the focal length of the measuring objective lens 11, d is the optical path from the reflective liquid crystal spatial light modulator 8 to the measuring objective lens 11, and λ is the wavelength of the light wave;

Fig. 2 is a phase grayscale diagram of the reflective liquid crystal spatial light modulator when the theoretical focusing distance x _F = 10 μm; Fig. 3 is a phase grayscale diagram of the reflective liquid crystal spatial light modulator when the theoretical focusing distance x _F = 50 μm.

Step 3. According to a series of theoretical focusing distances x _F selected in step 2, move the image sensor in the axial direction with a step size of 1 μm near the theoretical focusing distance x _F of the measurement objective lens 11 until the brightness of the spot reaches the maximum , the distance x' _F between the position of the image sensor and the focal point of the measuring objective lens 11 when the light spot brightness is maximum is the actual focusing distance;

_x'F are 5 μm, 10 μm, 17.5 μm, 25 μm, 27.5 μm, 35 μm, 37.5 μm, 42.5 μm, 47.5 μm, 55 μm, 80 μm, 105 μm, 205 μm and 340 μm;

The theoretical focusing distance x _F and the actual focusing distance x' _F and the error between them are shown in the table below.

Data comparison table of theoretical focusing distance x _F and actual focusing distance x' _F

The data in the table shows that within the range of 5-105 μm, the maximum error between the theoretical focusing distance x _F and the actual focusing distance x’ _F is not more than 5 μm, and when the focusing distance is above 100 μm, the theoretical focusing distance The error between the focal distance x _F and the actual focusing distance x' _F increases suddenly; it proves that when the allowable range of the focusing distance error of the device corresponding to this embodiment is below 5 μm, the focusing distance can reach 100 μm.

Step 4: Set the theoretical focusing distance x _F =0, configure the object to be scanned 10 as the object to be scanned, and adjust the object to be scanned to be at the focus position of the measurement objective lens 11; sequentially add the collection objective lens 13, the second pinhole 14, The three-dimensional precision displacement table 15 and the light intensity detector 16 make the light beam returning from the position of the object to be scanned along the original path pass through the measuring objective lens 11, transmit through the beam splitter 12, and be imaged by the collecting objective lens 13 to the second pinhole 14 position, and after passing through the second pinhole 14, it is imaged by the light intensity detector 16; a confocal axial scanning device based on a reflective liquid crystal spatial light modulator is formed, as shown in Figure 1;

Step 5. Keep the position of the object to be scanned in step 4. According to the actual required focusing distance x' _F , x' _F is 5μm, 10μm, 17.5μm, 25μm, 27.5μm, 35μm, 37.5μm, 42.5μm , 47.5μm, 55μm, 80μm and 105μm, through the theoretical focusing distance x _F corresponding to the actual focusing distance x' _F , the corresponding x _F are 5μm, 10μm, 15μm, 20μm, 25μm, 30μm, 35μm, 40μm , 45 μm, 50 μm, 75 μm, and 100 μm, the phase grayscale image of the reflective liquid crystal spatial light modulator is obtained as follows:

Wherein, _f0 is the focal length of the measuring objective lens 11, d is the optical path from the reflective liquid crystal spatial light modulator to the measuring objective lens 11, and λ is the wavelength of the light wave;

Step 6. According to the axial scanning depth of the shape of the object to be scanned, the phase grayscale image of the reflective liquid crystal spatial light modulator is changed, and at the same time, the light intensity detector 16 forms an image to obtain a confocal axial scanning curve. Among them, the comparison diagram of the confocal axial scanning curve when the focusing distance is 40 μm and when the focus is not adjusted is shown in Fig. 5 .

Using the confocal axial scanning device based on the reflective liquid crystal spatial light modulator of the present invention to realize the method for scanning the shape of the object, it includes the following specific steps:

Step 1. Set the theoretical focusing distance x _F = 0, configure the object to be scanned as the object to be scanned, and place the object to be scanned at a position where the distance from the measuring objective lens 11 is 2f ₀ ;

Step 2. Keep the relative position of the object to be scanned and the measuring objective lens 11 described in step 1 unchanged, and control the change of the phase grayscale map of the reflective liquid crystal spatial light modulator 8 through the computer, so that the theoretical adjustment corresponding to the phase grayscale map The focal distance x _F is from 0 μm to 105 μm in steps of 5 μm along the axial direction, and the relationship between the phase grayscale image and the theoretical focusing distance x _F is:

After adjusting the phase grayscale image of the reflective liquid crystal spatial light modulator 8 each time, the light intensity detector 16 is used for imaging to obtain the corresponding light intensity information;

Step 3. Obtain a corresponding confocal axial scanning curve according to each theoretical focusing distance x _F and light intensity information obtained in step 2;

Step 4: For a set of scanning curves obtained in step 3 with different axial depths, the maximum value of the ordinate in the connecting curves is integrated to obtain the shape of the object.

The horizontal distance between the maximum light intensity in the confocal axial scanning curve after focusing and the maximum light intensity without focusing is the focusing distance. If the maximum error between the focusing distance and the theoretical focusing distance x _F is within 5 μm, then It is considered that the x _F value at this time is an effective value;

Place the measured object at a distance of 2f ₀ from the measuring objective lens. Firstly, within the effective x _F range, for each corresponding grayscale image, a raster horizontal scan is performed on the measured object, so that the light intensity detector outputs a set of two-dimensional matrix values, and the gray values corresponding to different x _F values are continuously loaded. degree map, a group of two-dimensional light intensity matrices with different axial depths are obtained, and then the x _F value is used as the abscissa, and the corresponding element values of the same row and column in the matrix are drawn as a curve, and the maximum ordinate in the curve is found. The abscissa x _F value corresponding to the value is the shape of the object at this element, and finally the value corresponding to each element is integrated to obtain the shape of the object.

Claims (4)

1. The confocal axial scanning method based on the confocal axial scanning device of the reflective liquid crystal spatial light modulator, the confocal axial scanning device of the reflective liquid crystal spatial light modulator comprises a laser (1), a converging lens ( 2), the first pinhole (3), the illumination objective lens (4), the diaphragm (5), the polarizing beam splitter (6), the λ/4 wave plate (7), the reflective liquid crystal spatial light modulator (8), The measured object (10), the measuring objective lens (11), the beam splitter (12), the collecting objective lens (13), the second pinhole (14) and the light intensity detector (16), the parallel light beam emitted by the laser (1) passes through The converging lens (2) is imaged to the first pinhole (3), and the light beam passing through the first pinhole (3) is collimated by the illumination objective lens (4) and then enters the polarization beam splitter (6) in parallel, and the polarization beam splitter (6) ) divides the incident light into reflected light and transmitted light, the reflected light is S light, the transmitted light is P light, and the transmitted light reaches the reflective liquid crystal spatial light modulator (8) after passing through the λ/4 wave plate (7) ), the reflective liquid crystal spatial light modulator (8) totally reflects the arriving light, and the obtained total reflected light returns to the polarization beam splitter (6) according to the original optical path, and the reflected light reflected by the polarization beam splitter (6) enters the The beam splitter (12), the reflected light reflected by the beam splitter (12) is incident on the measurement objective lens (11), is imaged onto the measured object (10) through the measurement objective lens (11), and is reflected by the measured object (10) The reflected light returns to the beam splitter (12) along the original optical path, and the transmitted light transmitted by the beam splitter (12) is imaged by the collecting objective lens (13) to the position of the second pinhole (14), and passes through the second pinhole (14) incident to the photosensitive surface of the light intensity detector (16); It is characterized in that the method comprises specific steps as follows: Step 1. Set the theoretical focusing distance x _F = 0, configure the object to be scanned as the object to be scanned, and place the object to be scanned at a position where the distance from the measurement objective lens (11) is 2f ₀ ; Step 2. Keep the relative position of the object to be scanned and the measuring objective lens (11) described in step 1 unchanged, and control the phase grayscale change of the reflective liquid crystal spatial light modulator (8) by a computer, so that the phase grayscale The corresponding theoretical focusing distance x _F is stepped from 0 μm to 105 μm in the axial direction with a step size of 5 μm, and the relationship between the phase grayscale image and the theoretical focusing distance x _F is:

Wherein, _f0 is the focal length of the measuring objective lens, d is the optical path from the reflective liquid crystal spatial light modulator to the measuring objective lens, λ is the wavelength of the light wave, and ξ and η represent two-dimensional space coordinates; After adjusting the phase grayscale image of the reflective liquid crystal spatial light modulator (8) each time, the light intensity detector (16) is used for imaging to obtain corresponding light intensity information; Step 3: Obtain a corresponding confocal axial scanning curve according to each theoretical focusing distance x _F and light intensity information obtained in step 2. 2. the confocal axial scanning method based on the confocal axial scanning device of the reflective liquid crystal spatial light modulator according to claim 1, characterized in that the confocal axial scanning device of the reflective liquid crystal spatial light modulator It also includes a displacement platform (9), on which the measured object (10) is placed, and the displacement platform (9) can move one-dimensionally along the optical axis direction of the measuring objective lens (11). 3. the confocal axial scanning method based on the confocal axial scanning device of the reflective liquid crystal spatial light modulator according to claim 1, characterized in that the confocal axial scanning device of the reflective liquid crystal spatial light modulator It also includes a three-dimensional precision displacement platform (15), and a light intensity detector (16) is placed on the three-dimensional precision displacement platform (15). 4. the confocal axial scanning method based on the confocal axial scanning device of the reflective liquid crystal spatial light modulator according to claim 1, characterized in that the effective pixel area of the reflective liquid crystal spatial light modulator (8) It is 7.68mm×7.68mm, and the size of each pixel is 15μm×15μm.

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