Optical tweezers device for measuring elasticity of single cell

Abstract

The invention discloses an optical tweezers device for single cell elasticity measurement, which is characterized by comprising an optical tweezers module, a microscopic imaging module, a sample cell displacement module and a control and display module which are jointly positioned on a standard optical bread board, a laser device in the optical tweezers module emits a laser beam which sequentially passes through a laser beam expanding device, a power adjusting device, a first reflector, a beam coupling lens, a second reflector, a precise pinhole, a beam splitter prism and a rectangular prism reflector, and the laser beam is focused at an image space focal plane of a first objective lens to form an optical trap for capturing single cells. The single cell is controlled to perform relative motion at a constant speed through the control and display module and the sample cell displacement module, the fluid resistance generated by the motion causes the single cell to deform, and the microscopic imaging module receives image information and observes the deformation degree of the single cell to measure the elasticity of the single cell. The device has the advantages that the mobility is good, and the acting force for applying deformation is controllable.

Description

Technical Field

[0001] The present invention relates to the field of optical detection, in particular to an optical tweezers device for measuring the elasticity of a single cell. Background Art

[0002] The elasticity of single cells is an important parameter for measuring their deformability and mechanical properties, and is widely used in basic research and clinical diagnosis in the fields of hematology, tumors, and immunity. The measurement of single-cell elasticity is usually achieved by applying external stress to the single cell and observing the degree of deformation of the single cell. Optical tweezers technology has become an important means of applying single-cell deformation due to its advantages such as non-contact, high precision, and nanometer-level resolution. By utilizing the optical trapping force generated by a strongly focused laser beam, optical tweezers can achieve operations such as capturing, moving, and stretching single cells, thereby driving single cells to produce specific deformations.

[0003] Currently, most existing optical tweezers are large, desktop devices with bulky size, dispersed components, and poor mobility, limiting their widespread use in practical applications. Furthermore, existing optical tweezers struggle to deform single cells, lacking a simple and controllable solution for applying the force required to deform a single cell, making single-cell elasticity measurement difficult. Therefore, it is necessary to design an optical tweezers device for single-cell elasticity measurement to address these technical challenges. Summary of the Invention

[0004] (1) Technical problems solved

[0005] To address the shortcomings of existing technologies, the present invention provides an optical tweezers device for measuring single-cell elasticity. Through its integrated design and spatial layout, it addresses the bulky size and dispersed components of most existing optical tweezers devices, providing a viable solution for improving the device's mobility. Through optical path design and component optimization, it addresses the difficulties of existing optical tweezers in driving single-cell deformation and the difficulty in controlling the force exerted to apply deformation, thereby improving the operability of single-cell elasticity testing.

[0006] (2) Technical solution

[0007] In view of the above technical deficiencies, the purpose of the present invention is to provide an optical tweezers device for single-cell elasticity measurement, so as to solve the problems of poor mobility of most existing optical tweezers devices, difficulty in driving single-cell deformation, and difficulty in controlling the force exerted to deform single cells.

[0008] In order to achieve the above objectives, the present invention is implemented through the following technical solutions:

[0009] The present invention provides an optical tweezers device for single-cell elasticity measurement, comprising: an optical tweezers module, a microscopic imaging module, a sample cell displacement module, and a control and display module. These four modules are located on the same standard optical breadboard.

[0010] The optical tweezers module includes a laser device, a laser beam expander, a power adjustment device, a first reflector, a beam coupling lens, a second reflector, a precision pinhole, a beam splitter, a right-angle prism reflector, and a first objective lens; the microscopic imaging module includes an illumination light source, a second objective lens, the first objective lens, the right-angle prism reflector, the beam splitter, a filter, an eyepiece, and a CCD camera; the sample cell displacement module includes a sample cell, a stage, an X-axis manual micro-displacement stage, a Y-axis electric micro-displacement stage, a Z-axis manual micro-displacement stage, a motor control drive device, and a displacement joystick; the control and display module includes a small computer host and a small touch screen.

[0011] Preferably, the laser device includes a laser power supply and a laser emitter. The laser power supply is mounted on the standard optical breadboard. The laser emitter emits a laser light with a power of 200 mW, a wavelength of 532 nm, and an output spot diameter of 1.6 mm.

[0012] Preferably, the power adjustment device comprises a polarizer 1 and a polarizer 2. The operating wavelength of the polarizer 1 and the polarizer 2 is 400-700 nm.

[0013] Preferably, the laser beam expansion device includes a first lens and a second lens. The focal length of the first lens is -30 mm, the focal length of the second lens is 120 mm, the center distance between the first lens and the second lens is 90 mm, and the material of the first lens and the second lens is K9 glass.

[0014] Preferably, the focal length of the beam coupling lens is 175 mm, and the optical path length from the objective lens 1 is 335 mm. The beam coupling lens is made of K9 glass.

[0015] Preferably, the operating wavelength of the beam splitter prism is 400-700 nm, and the splitting ratio is 50:50.

[0016] Preferably, the precision pinhole has an aperture of 25 μm and an optical path length from the beam coupling lens to 175 mm.

[0017] Preferably, the first objective lens is an oil-based objective lens, placed inverted, with a focal length of 160 mm, a magnification of 100 times, and a numerical aperture of 1.2.

[0018] Preferably, the lighting source is fixed by an optical bracket at a height of 200 mm from the standard optical breadboard, with the light emitting surface facing downward. The lighting source is LED warm white light with a color temperature of 3000K.

[0019] Preferably, the objective lens 2 is placed upright, connected to the illumination light source, and facing the objective lens 1. The objective lens 2 has a magnification of 10 times and a numerical aperture of 0.25.

[0020] Preferably, the filter is located between the dichroic prism and the eyepiece, and has a central wavelength of 532 nm.

[0021] Preferably, the eyepiece is connected to the CCD camera, has a magnification of 0.5 times, and has an optical path length of 160 mm from the objective lens.

[0022] Preferably, the CCD camera has a resolution of 4608×3456 and a frame rate of 30 fps. The CCD camera is connected to the small computer host.

[0023] Preferably, the sample pool is placed above the stage, and the Z-axis manual micro-displacement stage, the Y-axis electric micro-displacement stage, the X-axis manual micro-displacement stage, the mounting base, and the standard optical breadboard are sequentially connected below.

[0024] Preferably, the motor control drive device includes a single-chip microcomputer development board fixed to the standard optical breadboard and a motor driver. The single-chip microcomputer development board is connected to the small computer host and the motor driver. The motor driver is connected to the Y-axis electric micro-displacement stage.

[0025] Preferably, the displacement rocker is fixed on the standard optical breadboard and connected to the single chip microcomputer development board.

[0026] Preferably, the small computer host is fixed on the standard optical breadboard, and its side interfaces are respectively connected to the CCD camera, the small touch display screen, and the motor control drive device.

[0027] Preferably, the small touch screen is clamped on an optical rod. The optical rod is fixed on the standard optical breadboard. The size of the small touch screen is 14 inches.

[0028] Preferably, the standard optical breadboard has two handles on either side and four silicone shock-absorbing wheels on the bottom. The standard optical breadboard measures 600mm x 300mm and has M6 threaded holes on the surface, with a spacing of 25mm between the holes. The standard optical breadboard is made of 6061-T6 aluminum alloy.

[0029] (3) Beneficial effects

[0030] The present invention provides an optical tweezers device for single cell elasticity measurement, which has the following beneficial effects:

[0031] This optical tweezers device for single-cell elasticity measurement integrates four modules: the optical tweezers module, the microscopy module, the sample cell displacement module, and the control and display module, using optimized optical path design and mechanical structure, onto a standard 600mm x 300mm optical breadboard. This device offers both comprehensive functionality and excellent portability.

[0032] 2. This optical tweezers device for single-cell elasticity measurement uses a low-power semiconductor laser with a power of 200mW and a wavelength of 532nm as its light source. The laser generates little heat during operation, eliminating the need for a bulky heat dissipation bellows. While enabling precise capture and manipulation of single cells, the device also saves space overall.

[0033] 3. This optical tweezers device for single-cell elasticity measurement is equipped with a Y-axis motorized micro-displacement stage, a motor driver, and a single-chip microcomputer development board. Operators can control the uniform motion of the Y-axis motorized micro-displacement stage by setting different movement speeds, thereby controlling the fluid resistance that causes single-cell deformation. This allows for simple and controllable force exerted on single cells, improving the operability of single-cell elasticity testing.

[0034] 4. This optical tweezers device for measuring single-cell elasticity features an integrated design, equipped with a small computer and touchscreen. This allows operators to observe single-cell deformation in real time, capturing and recording microscopic images of cells before and after deformation. Single-cell elasticity is assessed by measuring the length of single-cell morphologies and calculating relative deformation. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 It is a schematic structural diagram of the present invention as a whole;

[0036] Figure 2 Schematic diagram of the structure of the optical tweezers module of the present invention;

[0037] Figure 3 Schematic diagram of the structure of the microscopic imaging module of the present invention;

[0038] Figure 4 Schematic diagram of the structure of the sample cell displacement module of the present invention;

[0039] Figure 5 This is a schematic diagram of the structure of the control and display module of the present invention;

[0040] Figure 6 Schematic diagram of the optical path structure of the present invention;

[0041] Figure 7 This is a wiring diagram of the electronic circuit of the present invention.

[0042] In the figure: 101, standard optical breadboard; 102, silicone shock-absorbing wheel legs; 103, handle; 104, laser power supply; 105, lighting source regulator; 201, laser transmitter; 202, optical adjustment frame; 203, lens 1; 204, lens 2; 205, rotation adjustment frame; 206, polarizer 1; 207, polarizer 2; 208, reflector 1; 209, beam coupling lens; 210, optical cage system; 211, reflector 2; 212, precision pinhole; 213, beam splitter; 214, right-angle prism reflector; 215, objective lens 1; 3 01. Illumination light source; 302. Objective lens 2; 303. Filter; 304. Eyepiece; 305. CCD camera; 401. Sample cell; 402. Stage; 403. Z-axis manual micro-displacement stage; 404. X-axis manual micro-displacement stage; 405. Y-axis motorized micro-displacement stage; 406. Z-axis manual micro-displacement stage knob; 407. Micro-displacement stage base; 408. Displacement joystick; 409. X-axis manual micro-displacement stage knob; 410. Motor driver; 411. MCU development board; 501. Small touch screen; 502. Small computer host; 503. Optical extension rod; DETAILED DESCRIPTION

[0043] The present invention will be further described below in conjunction with the accompanying drawings and examples so that those skilled in the art can clearly understand the embodiments of the present invention and its technical effects. It should be noted that the following examples are only for illustrative description and do not limit the scope of protection of the present invention.

[0044] like Figures 1 to 7 As shown, the present invention provides an optical tweezers device for single-cell elasticity measurement, including an optical tweezers module, a microscopic imaging module, a sample pool displacement module, and a control and display module installed on a standard optical breadboard 101. The standard optical breadboard 101 has a specification of 300mm×600mm and is made of 6061-T6 aluminum alloy. The integrated lightweight design is convenient for movement and use in different scenarios. Two handles 103 are provided on both sides of the standard optical breadboard 101 for easy moving. Silicone shock-absorbing wheel legs 102 are installed at the bottom of the standard optical breadboard 101, which helps to isolate the adverse effects of external vibrations on the present invention and improve the stability of the operation of the present invention.

[0045] Specifically, the optical tweezers module comprises, in order of light propagation direction, a laser device, a laser beam expander, a power regulator, a reflector 1 208, a beam coupling lens 209, a reflector 211, a precision pinhole 212, a beam splitter 213, a right-angle prism reflector 214, and an objective lens 1 215. The laser device includes a laser power supply 104 and a laser emitter 201. The laser emitter 201 is secured within an optical adjustment mount 202. The laser emitter 201 is a semiconductor laser with a wavelength of 532 nm. The laser power supply 104 is secured to the standard optical breadboard 101 via four copper pillars. A switch is provided on the laser power supply 104 to facilitate turning the laser device on and off. The laser beam expander comprises a lens 1 203 and a lens 2 204, mounted within an optical cage system 210. The laser spot diameter can be adjusted by adjusting the focal length and position of the lens 1 203 and the lens 2 204. The laser beam expansion device has an expansion ratio of 1:4. The power adjustment device includes polarizer 1 206 and polarizer 2 207, which are fixed within a rotating adjustment frame 205. The rotating adjustment frame 205 is mounted within the optical cage system 210. By rotating the rotating adjustment frame 205, the angle between polarizer 1 206 and polarizer 207 is changed to adjust the output laser power. Reflector 1 208, reflector 2 211, and right-angle prism reflector 214 are mounted within the optical cage system 210. By adjusting the angles of reflector 1 208, reflector 2 211, and right-angle prism reflector 214, the direction of laser beam propagation is adjusted, optimizing the spatial structure of the optical path. The beam coupling lens 209 is mounted within the optical cage system 210. The focal length of the beam coupling lens 209 is 175 mm. The beam coupling lens 209 is used to adjust the radius of curvature of the laser Gaussian beam. The precision pinhole 212 is mounted within the optical cage system 210. The precision pinhole 212 serves to limit the laser pattern. The beam splitter prism 213 is mounted within the optical cage system 210. The beam splitter prism 213 reflects the laser beam and transmits the illumination beam, achieving a coaxial design of the optical tweezers and imaging optical paths. Objective lens 1 215 is fixed above the right-angle prism reflector 214. Objective lens 1 215 is positioned inverted to receive and focus the laser beam, forming a light trap for single-cell capture.

[0046] Specifically, the microscopic imaging module comprises, in order of light propagation direction, an illumination light source 301, objective lens 2 302, objective lens 1 215, right-angle prism reflector 214, dichroic prism 213, filter 303, eyepiece 304, and CCD camera 305. The illumination light source 301 is a warm white LED light source. The illumination light source 301 is equipped with an illumination light source regulator 105, which is fixed to the standard optical breadboard 101. The illumination light intensity is controlled by the illumination light source regulator 105 to meet actual lighting requirements. Objective lens 2 302 is placed upright, facing objective lens 1 215, and increases the illumination intensity by focusing the illumination light. The filter 303 is installed in the optical cage system 210. The filter 303 is a bandpass filter with a central wavelength of 532 nm to filter out the 532 nm laser beam and avoid the adverse effects of the laser beam on imaging. The eyepiece 304 is connected to the CCD camera 305 to collect the illumination light beam and image it on the CCD camera 305. The CCD camera 305 is installed in the optical cage system 210. The CCD camera 305 converts the optical signal of the single cell image information into an electrical signal and transmits it to the control and display module via a data line.

[0047] Specifically, the sample pool displacement module includes a sample pool 401, a stage 402, an X-axis manual micro-displacement stage 404, a Y-axis electric micro-displacement stage 405, a Z-axis manual micro-displacement stage 403, a motor control drive device, and a displacement rocker 408. The sample pool 401 is placed above the stage 402. The X-axis manual micro-displacement stage 404, the Y-axis electric micro-displacement stage 405, the Z-axis manual micro-displacement stage 403, and the stage 402 are connected from bottom to top. The micro-displacement stage base 407 is connected below the X-axis manual micro-displacement stage 404, and the micro-displacement stage base 407 is supported by a bracket and fixed on the standard optical breadboard 101. The movement of the X-axis manual micro-displacement stage 404 and the Z-axis manual micro-displacement stage 403 is controlled respectively by rotating the X-axis manual micro-displacement stage knob 409 and the Y-axis manual micro-displacement stage knob 406. The Y-axis electric micro-displacement stage 405 is controlled by the motor control drive device. The Y-axis electric micro-displacement stage 405 is controlled to move by a displacement command signal. The motor control drive device includes a single-chip microcomputer development board 411 and a motor driver 410 fixed to the standard optical breadboard 101. The motor control drive device is connected to a small computer host 502, the Y-axis electric micro-displacement stage 405, and the displacement rocker 408, respectively, to receive the displacement command signal and control the movement of the Y-axis electric micro-displacement stage 405. The displacement rocker 408 is fixed to the standard optical breadboard 101 and connected to the motor control drive device. The displacement command signal is issued by swinging the displacement rocker 408. After receiving the command signal, the single-chip microcomputer development board 411 controls the motor driver 410 to control the Y-axis electric micro-displacement stage 405 to move in a specified direction. The moving speed of the Y-axis electric micro-displacement stage 405 is adjusted by the numerical settings of the control and display module host computer.

[0048] Specifically, the control and display module includes the small computer host 502 and a small touch screen 501. The small computer host 502 is fixed to the upper right corner of the standard optical breadboard 101 to provide computing resources for the platform. Its side interfaces are respectively connected to the CCD camera 305, the small touch screen 501, and the motor control drive device for data transmission and processing. The small touch screen 501 is clamped on an optical connecting rod 503. The optical connecting rod 503 is fixed on the standard optical breadboard 101. The clamping height position and tilt angle of the small touch screen 501 are adjustable. The small touch screen 501 is connected to the small computer host 502 to display microscopic images and related calculation results in real time. The operator can control the movement of the Y-axis electric micro-displacement stage 405 through the touch interface of the small touch screen 501, capture and record the microscopic image information of the single cell before and after the deformation is applied, measure the morphological length and calculate the relative deformation amount to detect the elasticity of the single cell. The Y-axis electric micro-displacement stage 405 has the same moving speed, and the fluid resistance acting on the single cell is approximately equal. Under the action of equal fluid resistance, the larger the relative deformation of the single cell, the better the elasticity of the single cell.

[0049] Specifically, the sample pool displacement module and the control and display module electronic circuit wiring are as follows: Figure 7 As shown. The RS232 interface of the single-chip microcomputer development board 411 is connected to the USB interface of the small computer host 502. The single-chip microcomputer development board 411 is an Arduino MEGA-2560 model, powered by the USB interface of the small computer host 502, and burns code and sends control instructions through the RS232 interface. The DIR-, PUL-, and EN- pins on the motor driver 410 are connected to the GND pin on the single-chip microcomputer development board 411, and the EN+, DIR+, and PUL+ pins are respectively connected to the D5, D6, and D7 pins of the single-chip microcomputer development board 411. The single-chip microcomputer development board 411 can control the enable, direction, and pulse of the Y-axis electric micro-displacement stage 405 by giving different voltage levels to the D5, D6, and D7 pins. The A+, A-, B+, and B- pins on the motor driver 410 are respectively connected to the corresponding pins of the Y-axis electric micro-displacement stage 405, and the movement of the Y-axis electric micro-displacement stage 405 is controlled by four-step commutation. The VCC pin and GND pin of the displacement rocker 408 are connected to the +5V power supply pin and GND pin of the single-chip microcomputer development board 411, respectively, and are powered by the single-chip microcomputer development board 411. The VR pin of the displacement rocker 408 is connected to the A0 pin of the single-chip microcomputer development board 411, converting analog signals into digital signals and transmitting them to the single-chip microcomputer development board 411 to facilitate the control of the movement of the Y-axis electric micro-displacement stage 405.

[0050] When working, the specific steps of using the optical tweezers device for single cell elasticity measurement are as follows:

[0051] Step a. After turning on the laser device, the laser device emits a 532nm laser beam, which passes through the laser beam expansion device to increase the laser diameter so that the laser beam can fully fill the pupil at the objective lens 1 215. Then, the laser light power is adjusted by rotating the angle of the polarizer 2 207 through the power adjustment device. Then, the Gaussian beam curvature radius is adjusted through the beam coupling lens 209, and the laser mode is limited by the precision pinhole 212. In the optical path, the reflector 1 208, the reflector 2 211, the right-angle prism reflector 214 and the beam splitter prism 213 are used to adjust the direction of laser beam propagation and optimize the spatial layout of the optical path. Finally, the laser beam is focused at the objective lens 1 215 to form a light trap for single cell capture.

[0052] Step b. Turn on the illumination light source 301 and adjust the illumination light source regulator 105 to emit an illumination beam. The beam is focused by the second objective lens 302 to increase the illumination intensity. The light then passes through the first objective lens 215, the right-angle prism reflector 214, and the beam splitter 213. The light passes through the filter 303 to remove interference from the laser beam on the imaging process, and finally enters the eyepiece 304 and the CCD camera 305. The CCD camera 305 converts the optical signal into an electrical signal, which is then transmitted to the minicomputer via a data cable.

[0053] Step c. Turn on the small computer host 502 and the small touch screen 501 to display the image information obtained by the microscopic imaging module. Through the small touch screen 501, the operator can set parameters such as imaging exposure and adjust the position of the Z-axis manual micro-displacement stage 403 to ensure clear imaging.

[0054] Step d. Manually adjust the X-axis manual micro-displacement stage knob 409 and the Z-axis manual micro-displacement stage knob 406 to adjust the position of the stage 402 in the X-axis and Z-axis directions. The Y-axis motorized micro-displacement stage 405 is controlled by the displacement rocker 408 or the small touch screen 501 to adjust the position of the stage 402 in the Y-axis direction. Furthermore, by adjusting the position of the stage 402 in the X, Y, and Z-axis directions, the target single cell in the sample cell 401 is captured by the optical trap.

[0055] Step e. Set the movement speed of the Y-axis electric micro-displacement stage 405 through the small touch screen 501 (the speed setting range is 100μm / s to 300μm / s), and use the displacement rocker 408 or the small touch screen 501 to control the single cell captured in the light trap to move at a uniform speed in the Y-axis direction relative to the sample pool 401. The fluid resistance generated by the movement causes the single cell to deform. At the same movement speed, the fluid resistance that causes the deformation of the single cell is the same. By setting the movement speed of the Y-axis electric micro-displacement stage 405 to different sizes, the size of the fluid resistance that applies the deformation of the single cell can be controlled. The operator can observe the single cell deformation process in real time on the small touch screen 501, and capture and record the microscopic image information of the single cell before and after deformation. The elasticity of the single cell is detected by measuring the morphological length of the single cell and calculating the relative deformation amount. Under the action of the same size of fluid resistance, the larger the relative deformation amount of the single cell, the better the elasticity of the single cell.

[0056] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. An optical tweezers device for single cell elasticity measurement, comprising an optical tweezers module, a microscopic imaging module, a sample cell displacement module, and a control and display module; characterized in that: These four modules are located on the same standard optical breadboard; the optical tweezers module is composed of the following components in order of light propagation direction: laser device, laser beam expander, power adjustment device, reflector 1, beam coupling lens, reflector 2, precision pinhole, beam splitter, right-angle prism reflector, and objective lens 1; the sample cell displacement module includes a sample cell, a stage, an X-axis manual micro-displacement stage, a Y-axis electric micro-displacement stage, a Z-axis manual micro-displacement stage, a motor control drive device, and a displacement joystick; the control and display module includes a small computer host and a small touch screen.

2. The optical tweezers device for single cell elasticity measurement according to claim 1, characterized in that: The laser wavelength of the laser device is 532nm, the diameter of the output spot is 1.6mm, and the laser power is 200mW; the laser beam expansion device includes lens 1 and lens 2, the focal length of lens 1 is -30mm, the focal length of lens 2 is 120mm, and the distance between lens 1 and lens 2 is 90mm; the focal length of the beam coupling lens is 175mm, and the optical path length from the objective lens 1 is 335mm; the aperture of the precision pinhole is 25um, and the optical path length from the beam coupling lens is 175mm.

3. The optical tweezers device for single cell elasticity measurement according to claim 1, characterized in that: The small computer host and the small touch screen are fixed on the standard optical breadboard; the size of the small touch screen is 14 inches.

4. The optical tweezers device for single cell elasticity measurement according to claim 1, characterized in that: The standard optical breadboard has a size of 600mm×300mm. Two handles are provided on both sides of the standard optical breadboard, and four silicone shock-absorbing wheel legs are installed at the bottom.

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