A multi-scale photoacoustic microscopy method based on single-pixel imaging

Abstract

The present invention discloses a multi-scale photoacoustic microscopy method based on single-pixel imaging, comprising: 1) generating a series of stripe lights of different spatial frequencies in the XY two-dimensional plane to illuminate the sample, applying a phase shift to the stripe lights of each spatial frequency using a phase shifting method, and acquiring the photoacoustic signals at each phase shift using a single ultrasonic probe; 2) calculating Fourier spectrum coefficients based on the photoacoustic signals at each spatial frequency, modifying the Fourier spectrum coefficients by introducing the impulse response function of the ultrasonic probe, and then reconstructing the photoacoustic image based on the modified Fourier spectrum coefficients through an inverse Fourier transform based on a simplified point spread function (PSF); and 3) controlling the acquisition of the Fourier spectrum coefficients by controlling the spatial frequency of the stripe light, thereby achieving multi-scale imaging. The present method solves the problem that traditional photoacoustic microscopy, due to its fixed resolution, can only image at a single scale and cannot fully utilize multi-scale information.

Description

Technical Field

[0001] The present invention relates to the technical field of life science imaging, and in particular to a multi-scale photoacoustic microscopy method based on single-pixel imaging. Background Art

[0002] Photoacoustic imaging is a non-invasive imaging technology based on the photoacoustic effect that has emerged in the field of biomedical imaging in recent years. It combines the advantages of high contrast of optical imaging and high penetration of acoustic imaging, and has great clinical translation potential and application prospects.

[0003] Photoacoustic microscopy, a new hybrid imaging technology, combines the high contrast of optics with the low scattering and high resolution of acoustics, enabling deeper imaging of biological tissues. Currently, it has a wide range of applications in biological imaging, such as tumor detection, vascularization, and molecular imaging. However, traditional photoacoustic microscopy requires mechanical point scanning to acquire three-dimensional imaging information, resulting in slow imaging speeds. Furthermore, due to its fixed resolution, traditional photoacoustic microscopy can only image at a single scale and cannot fully utilize multi-scale information for integrated imaging. These factors limit the application of photoacoustic microscopy. Summary of the Invention

[0004] The purpose of the present invention is to address the shortcomings of the existing technology and provide a multi-scale photoacoustic microscopy imaging method based on single-pixel imaging to solve the problems of long imaging time and slow volume speed caused by ill-posed conditions (such as long mechanical point scanning time for volume imaging and fixed resolution).

[0005] In order to achieve the above-mentioned purpose, the present invention adopts the following technical solutions.

[0006] A multi-scale photoacoustic microscopy method based on single-pixel imaging comprises the following steps:

[0007] Step S1: Generate a series of stripe lights of different spatial frequencies in the XY two-dimensional plane to illuminate the sample using a spatial light modulator, apply a phase shift to the stripe lights of each spatial frequency using a phase shift method, and use a single ultrasonic probe to collect photoacoustic signals under each phase shift;

[0008] Step S2: Calculating Fourier spectrum coefficients based on the photoacoustic signal at each spatial frequency, modifying the Fourier spectrum coefficients by introducing the impulse response function of the ultrasound probe, and then reconstructing the photoacoustic image through inverse Fourier transform based on the modified Fourier spectrum coefficients and a simplified point spread function;

[0009] Step S3: Control the acquisition of Fourier spectrum coefficients by controlling the spatial frequency of the stripe light, thereby achieving multi-scale imaging.

[0010] Specifically, in step S1, a series of stripe lights with different spatial frequencies are generated in the XY two-dimensional plane by a spatial light modulator to illuminate the sample. and phase Stripe light, intensity distribution of stripe light Expressed as:

[0011] ;

[0012] In the above formula, x , y , z The positions of the illumination samples are xyz axis coordinates;

[0013] The phase shift method is used to apply phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect the photoacoustic signal under each phase shift. Expressed as:

[0014] ;

[0015] In the above formula, is the light absorption distribution, The stripe light irradiation area.

[0016] Specifically, the calculation method of the Fourier spectrum coefficient in step S2 is as follows:

[0017] The phase shift method is used to apply a phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect and obtain the photoacoustic signals of four phases, namely:

[0018] 、 、 、 ;

[0019] The light absorption distribution Fourier spectrum coefficient corresponding to the spatial frequency Expressed as:

[0020] ;

[0021] In the above formula, j Is an imaginary unit.

[0022] Furthermore, in step S2, the impulse response function of the ultrasound probe is introduced to correct the Fourier spectrum coefficients. Expressed as:

[0023] ;

[0024] In the above formula, β and is the nonlinear parameter that controls the attenuation rate of the photoacoustic signal. β and ; j is an imaginary unit; Photoacoustic signal k The amplitude of the resonance peak; Photoacoustic signal k The attenuation coefficient of the resonance peak; Photoacoustic signal k The time delay of a resonance peak; Photoacoustic signal k The phase of the resonance peak;

[0025] Corrected Fourier spectrum coefficients Expressed as:

[0026] ;

[0027] In the above formula, is the Wiener filter weight function, which is used to suppress the amplification of high-frequency noise;

[0028] ;

[0029] In the above formula, , is the conjugate impulse response function; is an empirical constant, , is the signal-to-noise ratio at the spatial frequency point.

[0030] Furthermore, in step S2, the photoacoustic image is reconstructed by inverse Fourier transform based on the simplified point spread function according to the corrected Fourier spectrum coefficients. Expressed as:

[0031] ;

[0032] In the above formula, In actual imaging systems, the spatial frequency allowed by the optical system is limited. For a given maximum spatial frequency of fringes, the objective lens with a given finite numerical aperture should satisfy ,in is the cutoff frequency of the objective lens, is the numerical aperture of the objective lens, is the wavelength of light; exp[*] is the exponential function; i is the imaginary unit; cc is the constant factor;

[0033] Ignoring constant terms and constant factors, the point spread function is further simplified in polar coordinates to:

[0034] ;

[0035] Photoacoustic image reconstructed by inverse Fourier transform based on simplified spread function PSF Expressed as:

[0036] .

[0037] Specifically, the control of the spatial frequency of the fringe light in step S3 is achieved by adjusting the wavelength of the light source, the fringe period, and the parameters of the projection system, and the Fourier spectrum coefficients are obtained based on a circular sampling strategy.

[0038] Furthermore, the circular sampling strategy defines a circular sampling area in the frequency domain, where the radius of the area corresponds to the required spatial frequency range; and obtains Fourier spectrum coefficients by uniformly sampling within the circular area.

[0039] Compared with the prior art, the present invention has the following beneficial effects:

[0040] 1. The method of the present invention uses stripe light of different spatial frequencies to illuminate the sample. The photoacoustic signal obtained by a single ultrasonic probe is used as the Fourier spectrum coefficient of the sample structure information at that spatial frequency. The image can be reconstructed by inverse Fourier transform. By controlling the spatial frequency of the stripes, the acquisition of the Fourier spectrum coefficient can be controlled, thereby achieving multi-scale imaging. This overcomes the defect of traditional photoacoustic microscopy that can only image at a single scale due to its fixed resolution and cannot fully utilize multi-scale information.

[0041] 2. Imaging experiments have verified that the method of the present invention can enhance the flexibility of photoacoustic imaging in actual imaging, expand the scope of application of photoacoustic imaging, and further enhance the integrated imaging capability of photoacoustic imaging through flexible and rapid multi-scale imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 This is a flow chart of a multi-scale photoacoustic microscopy method based on single-pixel imaging according to the present invention;

[0043] Figure 2 This is a schematic diagram of a multi-scale photoacoustic microscopy method based on single-pixel imaging according to the present invention;

[0044] Figure 3 This is a schematic diagram of a multi-scale photoacoustic imaging platform based on single-pixel imaging built using k-Wave;

[0045] Figure 4 It is the reconstruction result of multi-scale photoacoustic microscopy imaging of a single spherical absorber;

[0046] Figure 5 It is the reconstruction result of multi-scale photoacoustic microscopy imaging of multiple small spherical absorbers;

[0047] Figure 6 This is the result of reconstructing blood vessels using the reconstruction algorithm using the k-Wave: MATLAB simulation toolbox. DETAILED DESCRIPTION

[0048] In order to facilitate those skilled in the art to understand and implement the present invention, each step of the method proposed in the present invention is described in detail below. It should be understood that these embodiments are only used to illustrate the present invention and are not intended 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 claims appended hereto.

[0049] Example

[0050] like Figure 1 As shown, a multi-scale photoacoustic microscopy method based on single-pixel imaging includes the following steps:

[0051] Step S1: Generate a series of stripe lights of different spatial frequencies in the XY two-dimensional plane to illuminate the sample using a spatial light modulator, apply a phase shift to the stripe lights of each spatial frequency using a phase shift method, and use a single ultrasonic probe to collect photoacoustic signals under each phase shift;

[0052] Step S2: Calculating Fourier spectrum coefficients based on the photoacoustic signal at each spatial frequency, modifying the Fourier spectrum coefficients by introducing the impulse response function of the ultrasound probe, and then reconstructing the photoacoustic image through inverse Fourier transform based on the modified Fourier spectrum coefficients and a simplified point spread function;

[0053] Step S3: Control the acquisition of Fourier spectrum coefficients by controlling the spatial frequency of the stripe light, thereby achieving multi-scale imaging.

[0054] Specifically, in step S1, a series of stripe lights with different spatial frequencies are generated in the XY two-dimensional plane by a spatial light modulator to illuminate the sample. and phase Stripe light, intensity distribution of stripe light Expressed as:

[0055] ;

[0056] In the above formula, x , y , z The positions of the illumination samples are xyz axis coordinates;

[0057] The phase shift method is used to apply phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect the photoacoustic signal under each phase shift. Expressed as:

[0058] ;

[0059] In the above formula, is the light absorption distribution, The stripe light irradiation area.

[0060] Specifically, the calculation method of the Fourier spectrum coefficient in step S2 is as follows:

[0061] The phase shift method is used to apply a phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect and obtain the photoacoustic signals of four phases, namely:

[0062] 、 、 、 ;

[0063] The light absorption distribution Fourier spectrum coefficient corresponding to the spatial frequency Expressed as:

[0064] ;

[0065] In the above formula, j Is an imaginary unit.

[0066] Furthermore, in step S2, the impulse response function of the ultrasound probe is introduced to correct the Fourier spectrum coefficients. Expressed as:

[0067] ;

[0068] In the above formula, β and is the nonlinear parameter that controls the attenuation rate of the photoacoustic signal. β and ; j is an imaginary unit; Photoacoustic signal k The amplitude of the resonance peak; Photoacoustic signal k The attenuation coefficient of the resonance peak; Photoacoustic signal k The time delay of a resonance peak; Photoacoustic signal k The phase of the resonance peak;

[0069] Corrected Fourier spectrum coefficients Expressed as:

[0070] ;

[0071] In the above formula, is the Wiener filter weight function, which is used to suppress the amplification of high-frequency noise;

[0072] ;

[0073] In the above formula, , is the conjugate impulse response function; is an empirical constant, , is the signal-to-noise ratio at the spatial frequency point.

[0074] Furthermore, in step S2, the photoacoustic image is reconstructed by inverse Fourier transform based on the simplified point spread function according to the corrected Fourier spectrum coefficients. Expressed as:

[0075] ;

[0076] In the above formula, In actual imaging systems, the spatial frequency allowed by the optical system is limited. For a given maximum spatial frequency of fringes, the objective lens with a given finite numerical aperture should satisfy ,in is the cutoff frequency of the objective lens, is the numerical aperture of the objective lens, is the wavelength of light; exp[*] is the exponential function; i is the imaginary unit; cc is the constant factor;

[0077] Ignoring constant terms and constant factors, the point spread function is further simplified in polar coordinates to:

[0078] ;

[0079] Photoacoustic image reconstructed by inverse Fourier transform based on simplified spread function PSF Expressed as:

[0080] .

[0081] In this embodiment, the stripe light illumination effect of different spatial frequencies is equivalent to generating a virtual Bessel beam. The lateral resolution of the reconstructed photoacoustic image can be obtained from the full width at half maximum in the simplified point spread function expression: , by changing Different lateral resolutions can be obtained, achieving large-scale multi-scale imaging without moving. When the maximum spatial frequency of the stripe light is the cutoff frequency of the objective lens, that is, , the lateral resolution reaches the highest, ; Assume that the numerical aperture of the objective lens is ,wavelength , the lateral resolution is calculated to be approximately This lateral resolution can meet the high-resolution requirements of capillary imaging.

[0082] Specifically, the control of the spatial frequency of the fringe light in step S3 is achieved by adjusting the wavelength of the light source, the fringe period, and the parameters of the projection system, and the Fourier spectrum coefficients are obtained based on a circular sampling strategy.

[0083] Furthermore, the circular sampling strategy defines a circular sampling area in the frequency domain, where the radius of the area corresponds to the required spatial frequency range; and obtains Fourier spectrum coefficients by uniformly sampling within the circular area.

[0084] To verify the technical effect of the present invention, the multi-scale imaging capability was tested by using a single-pixel multi-scale imaging simulation model. The samples for this experimental verification were taken from a single spherical absorber, multiple spherical absorbers, and blood vessels, and different fringe spatial frequencies ( 、 、 ), most of the Fourier spectrum coefficients are obtained within the system's passband range. The feasibility and effectiveness of the method of the present invention are confirmed by comparing the resolution of the reconstructed images.

[0085] Figure 2 Schematic diagram of the multi-scale photoacoustic microscopy method based on single-pixel imaging of the present invention. Figure 2 (a) is a schematic diagram showing that a sequence of stripe light illumination is equivalent to generating a virtual Bessel spot (system point spread function). Figure 2 Middle (b) - Figure 2 (d) are respectively when the maximum spatial frequency of the stripe light is 、 、 The equivalent Bessel spot. Figure 2 (e) is Figure 2 Middle (b) - Figure 2 Middle (d) Light intensity distribution at the white dashed line, reflecting the resolution change; Figure 2 NI represents the normalized light intensity, and PSF represents the point spread function.

[0086] Figure 3Schematic diagram of a multi-scale photoacoustic imaging platform based on single-pixel imaging built using k-Wave. The experimental platform is configured with a 64-bit Intel® Core™ i7-8750H CPU @ 2.20 GHz and a Windows 10 operating system. The k-Wave simulation toolbox is used to build a multi-scale photoacoustic imaging platform based on single-pixel imaging. The k-Wave simulation toolbox is used to quickly solve a series of coupled first-order equations to simulate the propagation of photoacoustic signals. The three-dimensional acoustic medium space (including the perfectly matched layer PML used to meet the boundary conditions of forward transmission) has a pixel size of (At this point, the maximum fringe spatial frequency that the model can support is The sample can be set as needed and placed in a three-dimensional grid. The outer medium is water, the sound speed is set to 1500m / s, and the density is 1000kg / m 3 The number of ultrasonic transducers is 1, the center frequency is set to 50MHz, and the bandwidth is 80%. The axial distance between the ultrasonic transducer and the sample plane is Cosine fringe light of different spatial frequencies was generated according to the method of the present invention, and the Fourier spectrum was obtained using a circular sampling strategy. All simulations assumed an acoustically homogeneous medium with no absorption or scattering of sound.

[0087] like Figure 4 The figure shows the reconstruction result of multi-scale photoacoustic microscopy imaging of a single small spherical absorber. Figure 4 NPS in the equation stands for normalized power spectrum, and NPA stands for normalized photoacoustic amplitude. A small spherical absorber with a diameter of 2 microns is selected as the illumination sample. When the spatial frequency of the stripe light is When , that is, within the system passband allowable range, most of the Fourier spectrum coefficients are obtained, such as Figure 4 As shown in (a), at this time, the high-frequency information reflecting the detail information is obtained, and the reconstructed image is clearer; Figure 4 As shown in (b), the reconstructed ball outline is clearer. When, such as Figure 4 As shown in (c), at this time, some high-frequency information is not obtained, and the image details are lost; Figure 4 As shown in (d), the reconstructed image edges become less sharp. When, such as Figure 4 As shown in (e), the Fourier coefficients obtained at this time are basically low-frequency coefficients, the reconstructed image details are insufficient, and the lateral resolution is low, such as Figure 4 The Bessel side lobes shown in (f) are obvious (indicated by the white arrows). When, such as Figure 4 As shown in (g), only low-frequency Fourier coefficients are obtained at this time, and the resolution of the reconstructed ball is very poor, such as Figure 4 As shown in (h), the Bessel sidelobes are very obvious at this time (indicated by the white arrows).

[0088] The lateral resolution was quantitatively analyzed. Figure 4 The full width at half maximum (FWHM) of the photoacoustic signal distribution at the position of the white dotted line in the middle. Figure 4 As shown in (i), the changes in lateral resolution when the spatial frequency of the stripe light is different. 、 、 、 When, such as Figure 4 As shown in (j), the corresponding measured lateral resolutions are 、 、 、 ; The measured values ​​are consistent with the theoretical values, and the appropriate lateral resolution can be selected based on these data by quantitatively analyzing the lateral resolution.

[0089] like Figure 5 The figure shows the reconstruction results of multi-scale photoacoustic microscopy imaging of multiple small spherical absorbers. Figure 5 NPS in the equation represents the normalized power spectrum, and NPA represents the normalized photoacoustic amplitude. Four randomly distributed small spherical absorbers with a diameter of 2 μm were selected as the illumination sample. Figure 5 As shown in (a), when the spatial frequency of the fringe light is When the resolution is high, such as Figure 5 As shown in (b), the reconstructed spheres have clearer outlines and the four spheres can be clearly distinguished. Figure 5 As shown in (c), when the spatial frequency of the fringe light is When , the lateral resolution deteriorates by 2 times, but is still at the micron level, such as Figure 5 As shown in (d), the four balls can still be distinguished, but each ball becomes more blurred. Figure 5 As shown in (e), when the spatial frequency of the fringe light is When the lateral resolution deteriorates further by a factor of 2, reaching the level of more than ten microns, Figure 5 As shown in (f), the four small balls are difficult to distinguish.

[0090] The above details can be seen from the local magnification Figure 5 Medium (g) - Figure 5 This is more clearly seen in (i). Figure 5 (j) is Figure 5 Medium (g) - Figure 5The signal distribution at the white dotted line in (i) also clearly shows the change in resolution.

[0091] like Figure 6 The figure shows the result of blood vessel reconstruction using the reconstruction algorithm of k-Wave: MATLAB simulation toolbox. Figure 6 NPS represents the normalized power spectrum, and NPA represents the normalized photoacoustic amplitude. Large-scale multiscale imaging was performed using a virtual blood vessel. Simulation parameters were consistent with experimental performance tests. Figure 6 Middle (a) - Figure 6 (c) is the spatial frequency of the stripe light when the illumination 、 、 The Fourier spectrum obtained, and its corresponding reconstructed image is as follows Figure 6 Middle (d) - Figure 6 As shown in (f), it can be clearly seen that as the acquisition of high-frequency information increases, the blood vessels become clearer and clearer, and the edges of the blood vessels become more Rayleigh, reflecting the improvement of resolution. Figure 6 Medium (g) - Figure 6 (i) is Figure 6 Middle (b) - Figure 6 The partial enlarged image of the blood vessel in the white dotted frame in (f) is used to analyze the width of the blood vessel at the white dotted line. Figure 6 As shown in (j), it can be seen that when the spatial frequency of the fringe light is 、 、 When the measured widths are 、 、 The change in vessel width is consistent with the change in resolution. This indicates that the image reconstruction results of the present method are closer to the labeled image than those of traditional mechanical motion scanning methods and microprobe scanning methods. In other words, the present method has better imaging effects than traditional imaging methods.

[0092] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any other manner. Any person skilled in the art may utilize the above-disclosed technical content to modify or modify the present invention into equivalent embodiments. However, any simple modifications, equivalent variations, and modifications to the above embodiments that do not depart from the technical content of the present invention and are based on the technical essence of the present invention remain within the scope of protection of the present invention.

Claims

1. A multi-scale photoacoustic microscopy method based on single-pixel imaging, characterized in that: The following steps are involved: Step S1: Generate a series of stripe lights of different spatial frequencies in the XY two-dimensional plane to illuminate the sample using a spatial light modulator, apply a phase shift to the stripe lights of each spatial frequency using a phase shift method, and use a single ultrasonic probe to collect photoacoustic signals under each phase shift; Step S2: Calculating Fourier spectrum coefficients based on the photoacoustic signal at each spatial frequency, modifying the Fourier spectrum coefficients by introducing the impulse response function of the ultrasound probe, and then reconstructing the photoacoustic image through inverse Fourier transform based on the modified Fourier spectrum coefficients and a simplified point spread function; The calculation method of the Fourier spectrum coefficient is as follows: The phase shift method is used to apply a phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect and obtain the photoacoustic signals of four phases, namely: 、 、 、 ； The light absorption distribution Fourier spectrum coefficient corresponding to the spatial frequency Expressed as: ； In the above formula, j is an imaginary unit; The pulse response function of the ultrasonic probe is introduced to correct the Fourier spectrum coefficient. Expressed as: ； In the above formula, β and is the nonlinear parameter that controls the attenuation rate of the photoacoustic signal. β and ; j is an imaginary unit; Photoacoustic signal k The amplitude of the resonance peak; Photoacoustic signal k The attenuation coefficient of the resonance peak; Photoacoustic signal k The time delay of a resonance peak; Photoacoustic signal k The phase of the resonance peak; Corrected Fourier spectrum coefficients Expressed as: ； In the above formula, is the Wiener filter weight function, which is used to suppress the amplification of high-frequency noise; ； In the above formula, , is the conjugate impulse response function; is an empirical constant, , is the signal-to-noise ratio at the spatial frequency point; The point spread function Expressed as: ； In the above formula, In actual imaging systems, the spatial frequency allowed by the optical system is limited. For a given maximum spatial frequency of fringes, the objective lens with a given finite numerical aperture should satisfy ,in is the cutoff frequency of the objective lens, is the numerical aperture of the objective lens, is the wavelength of light; exp[*] is the exponential function; i is the imaginary unit; cc is the constant factor; Ignoring constant terms and constant factors, the point spread function is further simplified in polar coordinates to: ； Photoacoustic image reconstructed by inverse Fourier transform based on simplified spread function PSF Expressed as: ； Step S3: Control the acquisition of Fourier spectrum coefficients by controlling the spatial frequency of the stripe light, thereby achieving multi-scale imaging.

2. The multi-scale photoacoustic microscopy method based on single-pixel imaging according to claim 1, characterized in that: In step S1, a series of stripe lights with different spatial frequencies are generated in the XY two-dimensional plane by a spatial light modulator to illuminate the sample. and phase Stripe light, intensity distribution of stripe light Expressed as: ； In the above formula, x , y , z The positions of the illumination samples are xyz axis coordinates; The phase shift method is used to apply phase shift to the stripe light of each spatial frequency, and a single ultrasonic probe is used to collect the photoacoustic signal under each phase shift. Expressed as: ； In the above formula, is the light absorption distribution, The stripe light irradiation area.

3. The multi-scale photoacoustic microscopy method based on single-pixel imaging according to claim 1, characterized in that: In step S3, the control of the spatial frequency of the fringe light is achieved by adjusting the wavelength of the light source, the fringe period, and the parameters of the projection system, and the Fourier spectrum coefficients are obtained based on a circular sampling strategy.

4. The multi-scale photoacoustic microscopy method based on single-pixel imaging according to claim 3, characterized in that: The circular sampling strategy defines a circular sampling area in the frequency domain, where the radius of the area corresponds to the required spatial frequency range; and obtains Fourier spectrum coefficients by uniformly sampling within the circular area.

Images