Photoacoustic microscopic imaging axial resolution improving method based on single-pixel frequency domain acquisition

Abstract

The invention discloses a method for improving the axial resolution of photoacoustic microscopic imaging obtained based on a single-pixel frequency domain. The method comprises the following steps: 1) creating a Fourier substrate pattern with the spatial frequency of k in the axial direction; 2) respectively irradiating four Fourier substrate patterns with the spatial frequency of k and the initial phases of # imgabs0 #, # imgabs1 #, # imgabs2 # and # imgabs3 # to an imaging object, carrying out photoacoustic microscopic imaging, and acquiring data by using an ultrasonic transducer to obtain a response value of the ultrasonic transducer; 3) according to the obtained response value of the ultrasonic transducer, adopting a four-step phase shift method to obtain a Fourier coefficient corresponding to the spatial frequency k; and 4) gradually increasing the spatial frequency of the Fourier substrate pattern to the cut-off frequency of the objective lens, further obtaining Fourier coefficients corresponding to all spatial frequencies, obtaining a complete Fourier spectrum, and performing inverse Fourier transform to obtain a photoacoustic microscopic image after the axial resolution is increased. The method disclosed by the invention is expected to promote wider application of photoacoustic microscopic imaging in living cell and tissue imaging.

Description

Technical Field

[0001] The present invention relates to the technical field of photoacoustic imaging, and in particular to a method for improving the axial resolution of photoacoustic microscopic imaging based on single-pixel frequency domain acquisition. Background Art

[0002] Photoacoustic imaging is a new type of non-invasive, non-destructive imaging with the characteristics of high contrast and high resolution. After decades of research and development, the resolution of photoacoustic imaging has been continuously improved and has gradually entered the field of microscopy. In this process, a new type of photoacoustic imaging technology, namely photoacoustic microscopy (PAM), has emerged. PAM uses focused lasers to excite biological tissues to generate ultrasonic signals, and uses ultrasonic sensors to detect ultrasonic signals in a specific area to obtain sample structure information in that area. This technology can provide high-resolution, non-invasive deep imaging for biomedical research, and its lateral resolution can reach micrometers or even submicrometers. However, due to the constraints of the optical diffraction limit of the focusing objective and the acoustic diffraction limit of ultrasonic detection, the spatial resolution of PAM is difficult to further improve. In addition, the axial resolution of PAM is much lower than the lateral resolution, which results in the acquisition of three-dimensional structural information that cannot accurately reflect the true structure of biological tissues.

[0003] In order to further improve the axial resolution of PAM, it is crucial to obtain high-frequency spectrum information. Fourier single-pixel imaging provides an effective way to do this. Fourier single-pixel imaging technology can effectively restore the spectral information of the image. Integrating this technology into photoacoustic microscopy can make up for the shortcomings of photoacoustic microscopy in obtaining high-frequency signals and significantly improve the axial resolution. Summary of the invention

[0004] The purpose of the present invention is to address the problem that the axial resolution of photoacoustic microscopy is much smaller than the lateral resolution, and to provide a method for improving the axial resolution of photoacoustic microscopy based on single-pixel frequency domain acquisition, generate a sequence of structured light background patterns with different frequencies and phases in the axial direction, adopt a four-step phase shift method to gradually obtain the Fourier coefficients corresponding to each spatial frequency, obtain a complete Fourier spectrum, perform inverse Fourier transform on the obtained Fourier spectrum, and obtain a photoacoustic microscopy image with greatly improved axial resolution.

[0005] In order to achieve the above object, the present invention adopts the following technical solution.

[0006] A method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition comprises the following steps: Step S1, creating a Fourier basis pattern with an axial spatial frequency of k; Step S2: Set the four spatial frequencies to k and the initial phases to , , , The Fourier base patterns are respectively irradiated onto the imaging object to perform photoacoustic microscopy imaging, and the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer; Step S3, according to the obtained ultrasonic transducer response value, a four-step phase shift method is used to obtain the Fourier coefficient corresponding to the spatial frequency k; Step S4, gradually increase the spatial frequency of the Fourier basis pattern to the cutoff frequency of the objective lens, repeat steps S1 to S3, obtain the Fourier coefficients corresponding to all spatial frequencies within the cutoff frequency, obtain a complete Fourier spectrum, perform inverse Fourier transform on the obtained Fourier spectrum, and obtain a photoacoustic microscopy image with improved axial resolution.

[0007] Specifically, the spatial frequency k in step S1 is smaller than the bandwidth of the ultrasonic transducer.

[0008] Specifically, in step S1, a Fourier basis pattern with an axial spatial frequency of k is created, and the expression of the Fourier basis pattern is: ; In the above formula, is the average light intensity, is the contrast, is the Cartesian coordinate of the plane where the imaging object is located, is corresponding to The spatial frequency, It is the initial phase.

[0009] Specifically, in step S2, the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer. The imaging object is illuminated by the Fourier base pattern, and the signal intensity received by the ultrasonic transducer is for: ; In the above formula, is the illuminated area of ​​the Fourier basis pattern; is the reflection intensity; Corresponding to the response value of the ultrasonic transducer for: ; In the above formula, It is the response value caused by the noise signal received by the ultrasonic transducer; It is a constant factor, which is related to the magnification of the ultrasonic transducer and the spatial position relationship between the ultrasonic transducer and the object being measured.

[0010] Specifically, the four-step phase shift method described in step S3 is used to obtain the Fourier coefficient corresponding to the spatial frequency k, and the process is as follows: First, four samples with the same spatial frequency k and initial phases are , , , The Fourier basis patterns are irradiated onto the object respectively, and the four Fourier basis patterns are respectively , , , If it is expressed, then: ; Then, after trigonometric transformation, the above formula is rewritten into the following form: ; The response value of the ultrasonic transducer is further expressed as: ; Then the four-step phase shift algorithm is used to obtain the reflection intensity The Fourier transform integral of is: ; In the above formula, represents Fourier transform; represents an imaginary unit; Finally, the Fourier coefficient corresponding to the spatial frequency k is obtained : .

[0011] Specifically, the cutoff frequency of the objective lens in step S4 refers to the highest frequency boundary value of the signal allowed to pass through the objective lens used to excite the photoacoustic signal.

[0012] Compared with the prior art, the present invention has the following beneficial effects: The method of the present invention obtains the change of the detected object image in the Fourier spectrum domain by generating a sequence of structured light background patterns with different frequencies and phases in the axial direction, thereby avoiding the loss of the image Fourier spectrum information. At the same time, a four-step phase shift method is used to gradually obtain the Fourier coefficients corresponding to each spatial frequency, so that the Fourier spectrum of the image can be completely restored, and the obtained Fourier spectrum is inversely Fourier transformed, thereby reconstructing a photoacoustic microscopy image with greatly improved axial resolution.

[0013] The method of the present invention can obtain the ultra-high frequency photoacoustic signal spectrum of biological tissues through a low-frequency, low-cost ultrasonic probe, and is expected to promote the wider application of photoacoustic microscopy in living cell and tissue imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 This is a flow chart of a method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to the present invention; Figure 2 This is a comparative experimental result diagram of a rectangular sample in Example 1 of the present invention; Figure 3 is a comparative experimental result diagram of blood vessel samples in Example 2 of the present invention; Figure 4 This is a comparative experimental result diagram of the small ball sample in Example 3 of the present invention. DETAILED DESCRIPTION

[0015] In order to facilitate those of ordinary skill in the art to understand and implement the present invention, the following detailed description is given to each step of the method proposed by the present invention, and it should be understood that these embodiments are only used to illustrate the present invention and are not used to limit the scope of the present invention. In addition, it should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.

[0016] Example 1 like Figure 1 As shown, the present invention discloses a method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition, comprising the following steps: Step S1, creating a Fourier basis pattern with an axial spatial frequency of k; Step S2: Set the four spatial frequencies to k and the initial phases to , , , The Fourier base patterns are respectively irradiated onto the imaging object to perform photoacoustic microscopy imaging, and the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer; Step S3, according to the obtained ultrasonic transducer response value, a four-step phase shift method is used to obtain the Fourier coefficient corresponding to the spatial frequency k; Step S4, gradually increase the spatial frequency of the Fourier basis pattern to the cutoff frequency of the objective lens, repeat steps S1 to S3, obtain the Fourier coefficients corresponding to all spatial frequencies within the cutoff frequency, obtain a complete Fourier spectrum, perform inverse Fourier transform on the obtained Fourier spectrum, and obtain a photoacoustic microscopy image with improved axial resolution.

[0017] Specifically, the spatial frequency k in step S1 is smaller than the bandwidth of the ultrasonic transducer.

[0018] Specifically, in step S1, a Fourier basis pattern with an axial spatial frequency of k is created, and the expression of the Fourier basis pattern is: ;

[0019] In the above formula, is the average light intensity, is the contrast, is the Cartesian coordinate of the plane where the imaging object is located, is corresponding to The spatial frequency, It is the initial phase.

[0020] Specifically, in step S2, the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer. The imaging object is illuminated by the Fourier base pattern, and the signal intensity received by the ultrasonic transducer is for: ;

[0021] In the above formula, is the illuminated area of ​​the Fourier basis pattern; is the reflection intensity; Corresponding to the response value of the ultrasonic transducer for: ;

[0022] In the above formula, It is the response value caused by the noise signal received by the ultrasonic transducer; It is a constant factor, which is related to the magnification of the ultrasonic transducer and the spatial position relationship between the ultrasonic transducer and the object being measured.

[0023] Specifically, the four-step phase shift method described in step S3 is used to obtain the Fourier coefficient corresponding to the spatial frequency k, and the process is as follows: First, four samples with the same spatial frequency k and initial phases are , , , The Fourier basis patterns are irradiated onto the object respectively, and the four Fourier basis patterns are respectively , , , If it is expressed, then: ; Then, after trigonometric transformation, the above formula is rewritten into the following form: ; The response value of the ultrasonic transducer is further expressed as: ; Then the four-step phase shift algorithm is used to obtain the reflection intensity The Fourier transform integral of is: ; In the above formula, represents Fourier transform; represents an imaginary unit; Finally, the Fourier coefficient corresponding to the spatial frequency k is obtained : .

[0024] Specifically, the cutoff frequency of the objective lens in step S4 refers to the highest frequency boundary value of the signal allowed to pass through the objective lens used to excite the photoacoustic signal.

[0025] In this embodiment, Figure 2 The figure shows the photoacoustic microscopy imaging results of the method of the present invention and the traditional method for a rectangular sample. Figure 2 (a) is the original rectangular sample image. Figure 2 (b) is the photoacoustic microscopy imaging result of the rectangular sample by the method of the present invention. Figure 2 (c) shows the photoacoustic microscopy imaging result of a rectangular sample using the traditional method.

[0026] contrast Figure 2 (a) and Figure 2 As can be seen from (b), the reconstructed image of the photoacoustic microscopy imaging method of the present invention has a very clear outline, and the difference from the original image is almost only the amplitude; Figure 2 (a) and Figure 2 In (c), the imaging results of the two methods are significantly different. The reconstruction image of the traditional photoacoustic microscopy imaging can hardly distinguish the rectangular outline. Figure 2 (b) and Figure 2 By comparing with (c), it can be clearly seen that the method of the present invention improves the resolution.

[0027] In order to analyze the results more accurately, Figure 2 The data indicated by the arrow in (b) are fitted and analyzed, and the following is obtained: Figure 2 The data fitting diagram shown in (d) shows that the full width at half maximum (FWHM) of the photoacoustic microscopy imaging reconstruction image of the rectangular sample obtained by the method of the present invention is 0.73 μm. Figure 2 The data indicated by the arrow in (c) are fitted and analyzed, and the following is obtained: Figure 2 The data fitting diagram shown in (e) shows that the full width at half maximum (FWHM) of the conventional method for the reconstruction of the photoacoustic microscopy imaging of a rectangular sample is 35.09 μm. Comparing the results of the two methods, it can be concluded that the photoacoustic microscopy imaging method of the present invention greatly improves its axial resolution, proving the effectiveness of the present invention in improving the axial resolution of imaging.

[0028] Example 2 like Figure 3 The figure shows the photoacoustic microscopy imaging results of blood vessel samples using the method of the present invention and the traditional method. Figure 3 (a) is the original image of the blood vessel sample. Figure 3 (b) is the photoacoustic microscopy imaging result of the blood vessel sample by the method of the present invention. Figure 3 (c) shows the photoacoustic microscopy imaging result of vascular samples using traditional methods.

[0029] contrast Figure 3 (a) and Figure 3 As can be seen from (b), the reconstructed image of the photoacoustic microscopy imaging method of the present invention has a clearer outline and can completely reconstruct the vascular image; Figure 3 (a) and Figure 3 In (c), the imaging results of the two methods are significantly different. The reconstructed blood vessels of the traditional photoacoustic microscopy imaging method are blurred and incomplete. Figure 3 (b) and Figure 3 By comparing with (c), it can be clearly seen that the method of the present invention improves the resolution.

[0030] In order to analyze the results more accurately, Figure 3 The data indicated by the arrow in (b) are fitted and analyzed, and the following is obtained: Figure 3 The data fitting diagram shown in (d) shows that the full width at half maximum (FWHM) of the photoacoustic microscopy imaging reconstruction image of the blood vessel sample obtained by the method of the present invention is 1.8 μm. Figure 3 The data indicated by the arrow in (c) are fitted and analyzed, and the following is obtained: Figure 3 The data fitting diagram shown in (e) shows that the full width at half maximum (FWHM) of the conventional method for the reconstruction of the photoacoustic microscopy imaging of the vascular sample is 25μm. Comparing the results of the two, it can be concluded that the photoacoustic microscopy imaging method of the present invention greatly improves its axial resolution, proving the effectiveness of the present invention in improving the axial resolution of imaging.

[0031] Example 3 like Figure 4 The figure shows the photoacoustic microscopy imaging results of the sphere sample using the method of the present invention and the traditional method. The radius of the three spheres set in the sphere sample is 3 grids, the distance between sphere 1 and sphere 2 is 1 grid, and the distance between sphere 2 and sphere 3 is 10 grids. The centers of the three spheres are all located in the center column, so the center column is selected for axial resolution analysis. Since the distance between sphere 1 and sphere 2 is small, the center column data of sphere 1 and sphere 2 are selected for analysis here. Since the distance between sphere 1 and sphere 2 is close, when the image resolution is insufficient, the boundaries of sphere 1 and sphere 2 will be connected, and it is impossible to distinguish two independent spheres. Figure 4 (a) is the original image of the ball sample. Figure 4(b) is the photoacoustic microscopy imaging result of the small ball sample by the method of the present invention. Figure 4 (c) shows the photoacoustic microscopy imaging result of the small sphere sample using the traditional method.

[0032] according to Figure 4 In (b), it can be seen that the outlines of the three spheres are relatively clear, and sphere 1 and sphere 2 are not connected, indicating that the reconstructed image of the photoacoustic microscopy imaging method of the present invention has a high resolution. Figure 4 As can be seen in (c), ball 1 is indistinguishable from ball 2, and only ball 3, which is farther away, can be distinguished from ball 2, indicating that the reconstructed image resolution of traditional photoacoustic microscopy imaging is low.

[0033] In order to analyze the results more accurately, Figure 4 The data indicated by the arrow in (b) are fitted and analyzed, and the following is obtained: Figure 4 The data fitting diagram shown in (d) shows that the full width at half maximum (FWHM) of the photoacoustic microscopy imaging reconstruction image of the small ball sample by the method of the present invention is 3.66 μm; Figure 4 The data indicated by the arrow in (c) are fitted and analyzed, and the following is obtained: Figure 4 The data fitting diagram shown in (e) shows that the full width at half maximum (FWHM) of the photoacoustic microscopy imaging reconstruction image of the small ball sample by the traditional method is 83.17μm. Comparing the results of the two, it can be concluded that the photoacoustic microscopy imaging method of the present invention greatly improves its axial resolution, proving the effectiveness of the present invention in improving the axial resolution of imaging.

[0034] The above is only a preferred embodiment of the present invention, and does not limit the present invention in other forms. Any technician familiar with the profession may use the above disclosed technical content to change or modify it into an equivalent embodiment with equivalent changes. However, any simple modification, equivalent change and modification made to the above embodiment according to the technical essence of the present invention without departing from the technical solution of the present invention still belongs to the protection scope of the technical solution of the present invention.

Claims

1. A method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition, characterized in that: The following steps are involved: Step S1, creating a Fourier basis pattern with an axial spatial frequency of k; Step S2: Set the four spatial frequencies to k and the initial phases to , , , The Fourier base patterns are respectively irradiated onto the imaging object to perform photoacoustic microscopy imaging, and the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer; Step S3, according to the obtained ultrasonic transducer response value, a four-step phase shift method is used to obtain the Fourier coefficient corresponding to the spatial frequency k; Step S4, gradually increase the spatial frequency of the Fourier basis pattern to the cutoff frequency of the objective lens, repeat steps S1 to S3, obtain the Fourier coefficients corresponding to all spatial frequencies within the cutoff frequency, obtain a complete Fourier spectrum, perform inverse Fourier transform on the obtained Fourier spectrum, and obtain a photoacoustic microscopy image with improved axial resolution.

2. The method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to claim 1, characterized in that: In step S1, the spatial frequency k is smaller than the bandwidth of the ultrasonic transducer.

3. The method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to claim 2, characterized in that: In step S1, a Fourier basis pattern with an axial spatial frequency of k is created, and the expression of the Fourier basis pattern is: ； In the above formula, is the average light intensity, is the contrast, is the Cartesian coordinate of the plane where the imaging object is located, is corresponding to The spatial frequency, It is the initial phase.

4. The method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to claim 3, characterized in that: In step S2, the ultrasonic transducer is used to collect data to obtain the response value of the ultrasonic transducer. The imaging object is illuminated by the Fourier base pattern, and the signal intensity received by the ultrasonic transducer is for: ； In the above formula, is the illuminated area of ​​the Fourier basis pattern; is the reflection intensity; Corresponding to the response value of the ultrasonic transducer for: ； In the above formula, It is the response value caused by the noise signal received by the ultrasonic transducer; It is a constant factor, which is related to the magnification of the ultrasonic transducer and the spatial position relationship between the ultrasonic transducer and the object being measured.

5. The method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to claim 4, characterized in that: In step S3, the four-step phase shift method is used to obtain the Fourier coefficient corresponding to the spatial frequency k, and the process is as follows: First, four samples with the same spatial frequency k and initial phases are , , , The Fourier basis patterns are irradiated onto the object respectively, and the four Fourier basis patterns are respectively , , , If it is expressed, then: ； Then, after trigonometric transformation, the above formula is rewritten into the following form: ； The response value of the ultrasonic transducer is further expressed as: ； Then the four-step phase shift algorithm is used to obtain the reflection intensity The Fourier transform integral of is: ； In the above formula, represents Fourier transform; represents an imaginary unit; Finally, the Fourier coefficient corresponding to the spatial frequency k is obtained : 。 6. The method for improving the axial resolution of photoacoustic microscopy imaging based on single-pixel frequency domain acquisition according to claim 1, characterized in that: The cutoff frequency of the objective lens in step S4 refers to the maximum frequency boundary value of the signal allowed to pass through the objective lens used to excite the photoacoustic signal.

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