A dynamic quantitative differential interference contrast microscopy imaging system and method

Abstract

The present invention discloses a dynamic quantitative differential interference contrast microscopic imaging system and method. The system includes a differential interference contrast microscope, a Wollaston prism, a 4f imaging system, an analyzer, and a monochromatic black-and-white image sensor. An airspace carrier frequency quantitative differential phase imaging device composed of a Wollaston prism, a 4f imaging system, and an analyzer is connected to the output port on the side of the differential interference contrast microscope, so that two orthogonally polarized imaging light beams with a certain shear amount emitted from the differential interference contrast microscope interfere at the conjugate plane of the 4f imaging system to form an airspace carrier frequency differential interference pattern. Only one interference pattern is required to calculate the differential phase of the sample, and then the phase distribution of the sample is reconstructed by using the phase integration method according to the shear amount and shear direction of the differential interference contrast microscope. The present invention combines the high temporal phase stability provided by common-path interference and the low spatial phase noise generated by partially coherent light illumination, and has extremely high spatial phase sensitivity.

Description

Technical Field

[0001] The present invention relates to the technical field of optical microscopic measurement, and particularly relates to a dynamic quantitative differential interference contrast microscopic imaging system and method. Background Art

[0002] As a non-invasive imaging technology, optical microscopes play an important role in fields such as biomedical imaging. However, traditional bright-field microscopes can only obtain the amplitude information of the measured sample and cannot achieve high-contrast imaging. To solve this problem, specific imaging methods based on chemical or fluorescent dye labeling, namely fluorescence microscopes, have been proposed. They can detect the fluorescence signals generated when the dye is excited to achieve high-contrast imaging with molecular selectivity. However, this method is usually qualitative and dependent on sample preparation. In addition, photobleaching and phototoxicity caused by fluorescent labeling also limit the fluorescence imaging of living cells.

[0003] To achieve in-situ observation and research of living cells or tissues, label-free phase imaging technology is an effective imaging method, which can be divided into phase visualization and quantitative phase imaging. Among them, phase-contrast microscopes and differential interference contrast microscopes, as two typical phase imaging technologies, enable people to observe previously almost invisible unstained biological cells and tissues due to significant contrast gain. In contrast, quantitative phase imaging, as an imaging method that combines the characteristics of microscopes, holography, and light scattering technologies for phase recovery and optical metrology, is developing rapidly and has important application value in the field of life medicine research. Quantitative phase imaging can accurately measure the phase distribution in the phase-contrast image, thereby obtaining quantitative information on the morphology, internal structure, and refractive index distribution of biological cells and tissues, providing an important detection method for the life processes, disease diagnosis, and cell dynamics research of biological cells and tissues.

[0004] The differential interference contrast microscope obtains a differential interference pattern on the imaging plane through the common-path interference of two imaging light waves with orthogonal polarization directions and a small lateral shear. Using this method, the cause of the differential interference pattern is relatively clear, and it can be used for differential phase imaging of transparent samples and reflective samples. The phase image has the characteristics of high resolution and low noise, and can realize the optical sectioning function at different depths. However, the monochromatic shadow image can only qualitatively reflect the structure of the sample. Due to the non-linear relationship between intensity and phase, the acquired image cannot be directly used for quantitative calculation. In order to combine the above advantages of the differential interference contrast microscope and realize the quantitative phase imaging of the sample, differential phase reconstruction methods based on phase-shifting interference technology have been developed in the past, which can greatly improve the reconstruction accuracy of the sample phase. However, limited by the characteristics of the differential interference pattern (i.e., the background is relatively uniform, only showing high contrast at the sample, and there are no periodically distributed interference fringes), phase reconstruction methods with known phase shift amounts are usually required. This results in a single phase reconstruction method for differential interference patterns, strict control of phase shift accuracy, and difficulty in realizing dynamic measurement. Therefore, how to expand the phase reconstruction method for differential interference patterns, realize dynamic quantitative differential phase imaging, and thus realize quantitative phase imaging of label-free samples is a problem that needs to be solved. Summary of the Invention

[0005] In view of this, in order to solve the above problems in the prior art, the present invention proposes a dynamic quantitative differential interference contrast microscopy imaging system and method, which not only enables all high-precision phase-shifting algorithms based on unknown phase shift amounts to be applied to the phase reconstruction of differential interference patterns, but also can realize the dynamic quantitative phase imaging of differential interference contrast microscopes, and its time sampling rate is only limited by the camera acquisition rate.

[0006] The present invention solves the above problems through the following technical means:

[0007] On the one hand, the present invention provides a dynamic quantitative differential interference contrast microscopy imaging system, which sequentially includes along the optical path: an illumination module, a first polarizer, a first prism, a condenser, a stage, an imaging objective lens, a second prism, a tube lens, a third prism, a 4f imaging system, a second polarizer, and an image sensor;

[0008] The illumination module is used to emit illumination light;

[0009] The first polarizer is used to divide the illumination light emitted from the illumination module into linearly polarized illumination light waves along the x-axis and y-axis;

[0010] The first prism is used to cut the linearly polarized illumination light waves with polarization directions along the x-axis and y-axis at a small angle;

[0011] The condenser is used to collimate two orthogonally polarized light beams and irradiate the object to be measured at the stage;

[0012] The imaging objective lens is used to collect and converge the object light wave scattered by the object to be measured at the position of the second prism;

[0013] The second prism is used to combine two linearly polarized object light waves with a certain shear angle from the imaging objective lens;

[0014] The tube lens is used to converge two orthogonally polarized light beams and image them at the rear focal plane;

[0015] The third prism is used to deflect and separate the transmission directions of the object light waves from the tube lens with polarization directions along the x-axis and y-axis respectively;

[0016] The 4f imaging system is used to image two orthogonally polarized light beams and then enter the second polarizer;

[0017] The second polarizer is used to adjust the polarization states of two orthogonally polarized light beams and generate coherent superposition at the conjugate plane of the 4f imaging system to form a spatial frequency differential interference pattern;

[0018] The image sensor is used to obtain the spatial frequency differential interference pattern that can reflect the object to be measured, calculate the gradient distribution of the object to be measured using the Fourier transform algorithm, and finally reconstruct the phase distribution of the object to be measured using the phase integration method according to the gradient distribution of the object to be measured.

[0019] Preferably, the dynamic quantitative differential interference contrast microscopy imaging system further includes a mirror, and the mirror is located between the second prism and the tube lens.

[0020] Preferably, the illumination module is used to emit a monochromatic plane wave with a central wavelength of 633 nm and a spectral bandwidth of 5 nm; the magnification of the imaging objective lens is 20 times, and the numerical aperture is 0.45.

[0021] Preferably, the transmission axis of the first polarizer is in the x-z plane of the coordinate axis and the angle with the x-axis is 45°.

[0022] Preferably, the first prism is located at the front focal plane of the condenser, and its slow axis is in the x-z plane of the coordinate axis and parallel to the x-axis.

[0023] Preferably, the second prism is located at the rear focal plane of the imaging objective lens, and its slow axis is in the x-z plane of the coordinate axis and parallel to the x-axis.

[0024] Preferably, the third prism is located at the rear focal plane of the tube lens, and its slow axis is in the x-y plane of the coordinate axis and parallel to the x-axis.

[0025] Preferably, the 4f imaging system includes a first lens and a second lens, wherein the front focal plane of the first lens coincides with the rear focal plane of the tube lens, and the rear focal plane of the second lens is the conjugate plane of the 4f imaging system.

[0026] Preferably, the transmission axis of the second polarizer is in the x-y plane of the coordinate axes and forms an angle of 45° with the x-axis, allowing only the polarization components of the object light waves from the second lens and polarized in the x-axis and y-axis directions along the polarization direction of the second polarizer to pass through. Subsequently, coherent superposition occurs at the rear focal plane of the second lens to form a spatial frequency domain differential interference pattern; the image sensor is located at the conjugate plane of the 4f imaging system.

[0027] On the other hand, the present invention provides a method for dynamic quantitative differential interference contrast microscopy imaging, comprising the following steps:

[0028] Step 1: Turn on the illumination module. The monochromatic plane wave emitted by the illumination module is polarized by the first polarizer and then vertically incident on the first prism. After passing through the first prism, the monochromatic plane wave is sheared into two beams of light with a certain angle, and the polarization states of the two beams of light are orthogonal to each other; the two orthogonally polarized light beams are collimated by a condenser and then irradiated on the object to be measured at the stage; the object light waves scattered by the object to be measured are collected by the imaging objective lens and converged at the position of the second prism. At this time, the two orthogonally polarized light beams separated by the first prism are recombined by the second prism, and then reflected by a mirror and converged by the tube lens to form an image at the rear focal plane of the tube lens;

[0029] Step 2: After the two orthogonally polarized light beams converged by the tube lens pass through the third prism located at the rear focal plane of the tube lens, the transmission directions of the two orthogonally polarized light beams are deflected and separated. The two orthogonally polarized light beams separated by the third prism enter the second polarizer after passing through the 4f imaging system composed of the first lens and the second lens. After the polarization states of the two orthogonally polarized light beams are adjusted by the second polarizer, coherent superposition occurs at the conjugate plane of the 4f imaging system to form a spatial frequency domain differential interference pattern, which is then collected by the image sensor located at the conjugate plane of the 4f imaging system;

[0030] Step 3: After using the image sensor to obtain the spatial frequency domain differential interference pattern that can reflect the object to be measured, use the Fourier transform algorithm to calculate and obtain the gradient phase distribution of the object to be measured, and then reconstruct the phase distribution of the object to be measured according to the gradient phase distribution of the object to be measured.

[0031] Compared with the prior art, the beneficial effects of the present invention at least include:

[0032] (1). The dynamic quantitative differential interference contrast microscopy imaging system and method provided by the present invention encode the spatial carrier frequency into the differential interference pattern, thereby obtaining the carrier frequency differential interference pattern. Then, the differential phase of the sample to be measured is extracted from the carrier frequency differential interference pattern. Subsequently, according to the shear amount and shear direction of the differential interference contrast microscope, the phase distribution of the sample is reconstructed by phase integration. Different from the traditional phase reconstruction method using a known phase shift amount, not only can all high-precision phase shift algorithms based on an unknown phase shift amount be applied to the phase reconstruction of the differential interference pattern, thereby expanding the phase reconstruction method of the differential interference pattern, but also the dynamic quantitative phase imaging of the differential interference contrast microscope can be realized, and its time sampling rate is only limited by the camera acquisition rate.

[0033] (2). The dynamic quantitative differential interference contrast microscopy imaging system and method provided by the present invention have a simple structure and common-path interference, so that the air fluctuations and mechanical vibrations of the optical elements it receives are almost the same, and it has high stability. On the other hand, the present invention works well under partially coherent light illumination, so it has low spatial phase noise. In addition, it also has the advantages of no additional phase distortion, high light energy utilization rate, wide adaptability, etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following will briefly introduce the drawings required for the description of the embodiments. Obviously, the following drawings are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can also be obtained based on these drawings.

[0035] Figure 1 It is a structural diagram of a dynamic quantitative differential interference contrast microscopy imaging system proposed by the present invention;

[0036] Figure 2 It is for Figure 1 The spatial carrier frequency differential interference pattern of the airspace without a sample collected by the shown dynamic quantitative differential interference contrast microscopy imaging system;

[0037] Figure 3 It is for Figure 1 The spatial carrier frequency differential interference pattern of a polystyrene spherical cap collected by the shown dynamic quantitative differential interference contrast microscopy imaging system;

[0038] Figure 4 It is the differential phase of the polystyrene spherical cap reconstructed by the Fourier transform algorithm;

[0039] Figure 5 It is the differential phase of the polystyrene spherical cap reconstructed by the improved iterative algorithm (AIA);

[0040] Figure 6 It is for Figure 4 AndFigure 5 The difference in differential phase of the polystyrene spherical cap;

[0041] Figure 7 The phase distribution of the polystyrene spherical cap is reconstructed by using the phase integration method according to the shear amount and shear direction of the differential interference contrast microscope;

[0042] Figure 8 The phase profile of the polystyrene spherical cap reconstructed by the system and method provided in the embodiment of the present invention and the comparison diagram of the phase profile reconstructed by the digital holographic microscope.

[0043] Explanation of reference numerals:

[0044] 101. LED illumination module; 102. First polarizer; 103. First Nomarski prism; 104. Condenser; 105. Stage; 106. Infinity imaging objective; 107. Second Nomarski prism; 108. Mirror; 109. Tube lens; 110. Wollaston prism; 111. First lens; 112. Second lens; 113. Second polarizer; 114. Monochromatic black and white image sensor. Detailed implementation manners

[0045] In order to make the above objects, features and advantages of the present invention more obvious and understandable, the technical solutions of the present invention will be described in detail below with reference to the drawings and specific embodiments. It should be noted that the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts belong to the scope of protection of the present invention.

[0046] Embodiment 1

[0047] Refer to Figure 1 , a dynamic quantitative differential interference contrast microscopy imaging system provided by an embodiment of the present invention sequentially includes along the optical path: an LED illumination module 101, a first polarizer 102, a first Nomarski prism 103, a condenser 104, a stage 105, an infinity imaging objective 106, a second Nomarski prism 107, a mirror 108, a tube lens 109, a Wollaston prism 110, a first lens 111, a second lens 112, a second polarizer 113 and a monochromatic black and white image sensor 114.

[0048] The LED illumination module 101 is used to emit a monochromatic plane wave with a central wavelength of 633 nm and a spectral bandwidth of 5 nm.

[0049] The transmission axis of the first polarizer 102 is in the x-z plane of the illustrated coordinate axes and makes an angle of 45° with the x-axis, and is used to divide the illumination light emitted from the LED illumination module 101 into linearly polarized lights along the x-axis and the y-axis.

[0050] The first Nomarski prism 103 is located at the front focal plane of the condenser lens 104. Its slow axis is in the x-z plane of the illustrated coordinate axes and is parallel to the x-axis, and is used to shear the linearly polarized illumination light waves with polarization directions along the x-axis and the y-axis respectively by a small angle.

[0051] The condenser lens 104 is used to collimate the two orthogonally polarized lights and irradiate them on the object to be measured at the stage 105.

[0052] The magnification of the infinity imaging objective lens 106 is 20 times and the numerical aperture is 0.45. It is used to collect and converge the object light waves scattered by the object to be measured at the position of the second Nomarski prism 107.

[0053] The second Nomarski prism 107 is located at the rear focal plane of the infinity imaging objective lens 106. Its slow axis is in the x-z plane of the illustrated coordinate axes and is parallel to the x-axis, and is used to combine the two linearly polarized object light waves with a certain shear angle from the infinity imaging objective lens 106.

[0054] The tube lens 109 is used to converge the two orthogonally polarized lights reflected by the mirror 108 and form an image at the rear focal plane.

[0055] The Wollaston prism 110 is located at the rear focal plane of the tube lens 109. Its slow axis is in the x-y plane of the illustrated coordinate axes and is parallel to the x-axis, and is used to deflect the transmission directions of the object light waves with polarization directions along the x-axis and the y-axis from the tube lens 109 and separate them, and the separation angle is 1°.

[0056] The first lens 111 and the second lens 112 form a 4f imaging system with a magnification of 1 time. The front focal plane of the first lens 111 coincides with the rear focal plane of the tube lens 109, the rear focal plane of the second lens 112 is the conjugate plane of the 4f imaging system, and the first lens 111 and the second lens 112 are achromatic doublet lenses with a focal length of 80 mm each. The 4f imaging system is used to image the two orthogonally polarized lights and then enter the second polarizer 113.

[0057] The transmission axis of the second polarizer 113 is in the x-y plane of the illustrated coordinate axes and makes an angle of 45° with the x-axis, and only allows the polarization components along the polarization direction of the second polarizer 113 in the object light waves with polarization directions along the x-axis and the y-axis from the second lens 112 to pass through. Subsequently, coherent superposition occurs at the rear focal plane of the second lens 112 and an in-air carrier frequency differential interference pattern is formed.

[0058] The monochromatic black-and-white image sensor 114 is located at the conjugate plane of the 4f imaging system. After obtaining the spatial frequency differential interference pattern of the object to be measured, the Fourier transform algorithm is used to calculate the gradient distribution of the object to be measured. Finally, based on the gradient distribution of the object to be measured, the phase distribution of the object to be measured is reconstructed by the phase integration method.

[0059] Example 2

[0060] The present invention provides a dynamic quantitative differential interference contrast microscopy imaging method, including the following steps:

[0061] S1. The monochromatic plane wave emitted by the LED illumination module 101 is vertically incident on the first Nomarski prism 103 after polarization by the first polarizer 102. After passing through the first Nomarski prism 103, the monochromatic plane wave is sheared into two beams of light with a certain angle, and the polarization states of the two beams of light are orthogonal to each other. The two orthogonally polarized light beams are collimated by the condenser lens 104 and then irradiated on the object to be measured at the stage 105. The object wave scattered by the object is collected by the infinity imaging objective lens 106 and converged at the position of the second Nomarski prism 107. At this time, the two orthogonally polarized light beams separated by the first Nomarski prism 103 are recombined by the second Nomarski prism 107, and then reflected by the mirror 108 and converged by the tube lens 109 and imaged at the rear focal plane of the tube lens 109.

[0062] S2. After the two orthogonally polarized light beams converged by the tube lens 109 pass through the Wollaston prism 110 located at the rear focal plane of the tube lens 109, the transmission directions of the two orthogonally polarized light beams are deflected and separated, and the separation angle is 1°. Next, the two orthogonally polarized light beams enter the second polarizer 113 after passing through the 4f imaging system composed of the first lens 111 and the second lens 112. After the polarization states of the two orthogonally polarized light beams are adjusted by the second polarizer 113, coherent superposition occurs at the conjugate plane of the 4f imaging system to form a spatial frequency differential interference pattern, and finally, it is collected by the monochromatic black-and-white image sensor 114 located at the conjugate plane of the 4f imaging system.

[0063] S3. After using the monochromatic black-and-white image sensor 114 to obtain the spatial frequency differential interference pattern of the object to be measured, the Fourier transform algorithm is used to calculate the gradient distribution of the object to be measured. Finally, based on the gradient distribution of the object to be measured, the phase distribution of the object to be measured is reconstructed by the phase integration method.

[0064] Experimental results

[0065] To test a dynamic quantitative differential interference contrast microscopy imaging system and method provided by the present invention, an experimental optical path provided by an embodiment of the present invention was built, and quantitative phase imaging was performed on a polystyrene spherical cap. During measurement, the LED illumination module 101 was turned on, and a spatial carrier differential interference pattern without a sample was collected by the monochromatic black-and-white image sensor 114 as shown in Figure 2 . The phase distribution of the spatial carrier differential interference pattern without a sample was calculated using the Fourier transform algorithm. Then, the prepared polystyrene spherical cap was placed on the stage 105, and a spatial carrier differential interference pattern with the sample was collected as shown in Figure 3 . After reconstructing its phase using the Fourier transform algorithm, the phase distribution of the spatial carrier differential interference pattern without a sample was subtracted to obtain the differential phase of the polystyrene spherical cap as shown in Figure 4 . Next, 200 spatial carrier differential interference patterns with random phase shifts were further collected, and the differential phase of the spherical cap was extracted using the improved iterative algorithm (AIA) (please refer to Figure 5 ) for comparison. The result of subtracting the differential phases calculated by the two methods is shown in Figure 6 . It can be seen that the differential phases reconstructed by the two algorithms are almost the same. Finally, according to the shear amount and shear direction of the differential interference contrast microscope, the phase of the sample was reconstructed using the phase integration method as shown in Figure 7 . To test the measurement accuracy of the proposed method, we used a digital holographic microscope to measure the same polystyrene spherical cap. The cross-section of the phase distribution of the sample obtained by the digital holographic microscope was compared with the phase profile reconstructed by the system and method provided by the embodiment of the present invention (please refer to the position of the white dotted line in Figure 7 ), and the result is shown in Figure 8 . As can be seen from Figure 8 , the phase reconstructed by the system and method provided by the embodiment of the present invention is consistent with the result obtained by the digital holographic microscope.

[0066] In summary, the system of the present invention is composed of a differential interference contrast microscope, a Wollaston prism, a 4f imaging system, an analyzer, and a monochromatic black-and-white image sensor. An airspace carrier quantitative differential phase imaging device composed of a Wollaston prism, a 4f imaging system, and an analyzer is connected to the output port on the side of the differential interference contrast microscope, so that two orthogonally polarized imaging light beams with a certain shear amount emitted from the differential interference contrast microscope interfere at the conjugate plane of the 4f imaging system to form an airspace carrier differential interference pattern. Only one interference pattern is required to calculate the differential phase of the sample, and then the phase distribution of the sample is reconstructed using the phase integration method according to the shear amount and shear direction of the differential interference contrast microscope. The present invention combines the high temporal phase stability provided by common-path interference and the low spatial phase noise generated by partially coherent light illumination, making it have extremely high spatial phase sensitivity.

[0067] A dynamic quantitative differential interference contrast microscopy imaging system and method provided by the present invention can perform rapid and high-precision quantitative phase imaging on unstained samples. Different from traditional phase reconstruction methods using known phase shift amounts, it not only enables all high-precision phase shift algorithms based on unknown phase shift amounts to be applied to the phase reconstruction of differential interference patterns, thereby expanding the phase reconstruction methods of differential interference patterns, but also can achieve dynamic quantitative phase imaging of differential interference contrast microscopes, and its time sampling rate is only limited by the camera acquisition rate. The present invention combines the high temporal phase stability provided by common-path interference and the low spatial phase noise generated by partially coherent light illumination, making it have extremely high spatial phase sensitivity. In addition, the present invention also has the advantages of simple structure, no additional phase distortion, high light energy utilization rate, wide adaptability, etc., and can achieve high-precision dynamic quantitative imaging of differential phases, which can provide an effective means for quantitative measurement and observation in biomedicine.

[0068] The above-described embodiments merely represent several implementation manners of the present invention, and the description thereof is relatively specific and detailed, but it should not be construed as a limitation to the scope of the patent of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the appended claims.

Claims

1. A dynamic quantitative differential interference contrast microscopy imaging system, characterized in that It sequentially includes along the optical path: an illumination module, a first polarizer, a first prism, a condenser lens, a stage, an imaging objective lens, a second prism, a tube lens, a third prism, a 4f imaging system, a second polarizer, and an image sensor; The illumination module is used to emit illumination light; The first polarizer is used to divide the illumination light emitted from the illumination module into linearly polarized illumination light waves along the x-axis and the y-axis; The first prism is used to cut the linearly polarized illumination light waves with polarization directions along the x-axis and the y-axis at a small angle; The condenser lens is used to collimate two orthogonally polarized light beams and irradiate them on the object to be measured at the stage; The imaging objective lens is used to collect and converge the object light waves scattered by the object to be measured at the position of the second prism; The second prism is used to combine two linearly polarized object light waves with a certain shear angle from the imaging objective lens; The tube lens is used to converge two orthogonally polarized light beams and image them at the rear focal plane; The third prism is used to deflect and separate the propagation directions of the object light waves with polarization directions along the x-axis and the y-axis from the tube lens; the third prism is located at the rear focal plane of the tube lens, and its slow axis is in the x-y plane of the coordinate axes and parallel to the x-axis; The 4f imaging system is used to image two orthogonally polarized light beams and then enter the second polarizer; the 4f imaging system includes a first lens and a second lens, where the front focal plane of the first lens coincides with the rear focal plane of the tube lens, and the rear focal plane of the second lens is the conjugate plane of the 4f imaging system; The second polarizer is used to adjust the polarization states of two orthogonally polarized light beams and generate coherent superposition at the conjugate plane of the 4f imaging system to form a spatial frequency domain differential interference pattern; The image sensor is used to obtain a spatial frequency domain differential interference pattern that can reflect the object to be measured, calculate the gradient distribution of the object to be measured using the Fourier transform algorithm, and finally reconstruct the phase distribution of the object to be measured using the phase integration method according to the gradient distribution of the object to be measured; The transmission axis of the second polarizer is in the x-y plane of the coordinate axes and forms an angle of 45° with the x-axis, and only allows the polarization components of the object light waves with polarization directions along the x-axis and the y-axis from the second lens along the polarization direction of the second polarizer to pass through, and then generate coherent superposition at the rear focal plane of the second lens to form a spatial frequency domain differential interference pattern; the image sensor is located at the conjugate plane of the 4f imaging system.

2. The dynamic quantitative differential interference contrast microscopy imaging system according to claim 1, wherein The dynamic quantitative differential interference contrast microscopy imaging system further includes a reflector, and the reflector is located between the second prism and the tube lens.

3. The dynamic quantitative differential interference contrast microscopy imaging system according to claim 1, wherein The illumination module is used to emit a monochromatic plane wave with a central wavelength of 633 nm and a spectral bandwidth of 5 nm; the magnification of the imaging objective lens is 20 times, and the numerical aperture is 0.

45.

4. The dynamic quantitative differential interference contrast microscopy imaging system according to claim 1, characterized in that, The transmission axis of the first polarizer is in the x-z plane of the coordinate axes and forms an angle of 45° with the x-axis.

5. The dynamic quantitative differential interference contrast microscopy imaging system according to claim 1, wherein The first prism is located at the front focal plane of the condenser lens, and its slow axis is in the x-z plane of the coordinate axes and parallel to the x-axis.

6. The dynamic quantitative differential interference contrast microscopy imaging system according to claim 1, wherein The second prism is located at the rear focal plane of the imaging objective lens, and its slow axis is in the x-z plane of the coordinate axes and parallel to the x-axis.

7. A dynamic quantitative differential interference contrast microscopy imaging method, characterized in that, It includes the following steps: Step 1: Turn on the illumination module. The monochromatic plane wave emitted by the illumination module is vertically incident on the first prism after polarization by the first polarizer. After passing through the first prism, the monochromatic plane wave is sheared into two beams of light with a certain angle, and the polarization states of the two beams of light are orthogonal to each other. The two orthogonally polarized light beams are collimated by a condenser lens and then irradiated on the object to be measured at the stage. The object wave scattered by the object to be measured is collected by the imaging objective lens and converged at the position of the second prism. At this time, the two orthogonally polarized light beams separated by the first prism are recombined by the second prism, and then reflected by a mirror and converged by the tube lens to form an image at the rear focal plane of the tube lens. Step 2: After the two orthogonally polarized light beams converged by the tube lens pass through the third prism located at the rear focal plane of the tube lens, the transmission directions of the two orthogonally polarized light beams are deflected and separated. The two orthogonally polarized light beams separated by the third prism enter the second polarizer after passing through the 4f imaging system composed of the first lens and the second lens. After the polarization states of the two orthogonally polarized light beams are adjusted by the second polarizer, they produce coherent superposition at the conjugate plane of the 4f imaging system to form a spatial frequency differential interference pattern, which is then collected by the image sensor located at the conjugate plane of the 4f imaging system. Step 3: After using the image sensor to obtain the spatial frequency differential interference pattern that can reflect the object to be measured, use the Fourier transform algorithm to calculate and obtain the gradient phase distribution of the object to be measured, and then reconstruct the phase distribution of the object to be measured according to the gradient phase distribution of the object to be measured.

Images