CN112509122A - Method and device for detecting concentration and activity of high-concentration microparticles based on multi-height lensless digital holography - Google Patents
Method and device for detecting concentration and activity of high-concentration microparticles based on multi-height lensless digital holography Download PDFInfo
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Abstract
The invention provides a method and a device for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography, which comprises the following steps: acquiring original holograms of a plurality of sub-pixel displacements with different z-axis distances in a sample to be detected; performing sub-pixel registration processing on the obtained original hologram; based on the original hologram after the sub-pixel registration processing, restoring the amplitude and phase information of the image by adopting a single-beam multi-intensity reconstruction algorithm; performing super-resolution processing on the original hologram by adopting a convex set projection iterative algorithm, and improving the resolution of the hologram; performing three-dimensional reconstruction on the super-resolution image to obtain the length, width and thickness information of the sample to be detected, and realizing the detection of the activity of the sample to be detected; and calculating the refractive index of the sample to be detected based on the phase information of the hologram after the super-resolution processing through the relationship between the concentration of each high-concentration microparticle and the refractive index obtained in advance, thereby obtaining the concentration of the sample to be detected and realizing the detection of the concentration of the high-concentration microparticle.
Description
Technical Field
The invention relates to the technical field of high-concentration microparticle concentration and activity detection, in particular to a method and a device for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography.
Background
The micro-particles are widely present in natural environments and medical fields, such as marine micro-plastics, microalgae cells in ship ballast water and cancer cells in the medical field, once a large amount of the micro-plastics are present in the sea, the original ecological balance of the sea is destroyed, and even the micro-plastics threaten the life health of other organisms and human bodies in the sea, once the microalgae cells in the ship ballast water are discharged into the sea, the micro-algae cells have extremely strong environmental adaptation capability and cover the surface of a water body to cause the phenomenon that fishes in the sea die due to oxygen deficiency, and the cancer cells in the medical field damage the life health of human bodies, so that the detection of the micro-particles has important significance for the natural environments and the medical field.
The existing detection methods of the micro-plastics are microscopy, infrared spectroscopy, Raman spectroscopy, scanning electron microscopy and the like, the detection methods of the micro-algae cells mainly comprise optical microscopy, fluorescence microscopy, flow cytometry, coulter counting, image analysis, molecular and biochemical methods and the like, in the medical field, the cancer cells are the root cause of cancer, the earlier the detection of the cancer cells causes less harm to human bodies, and the traditional detection methods of the lung cancer cells mainly comprise the following methods: x-ray examination, percutaneous lung biopsy, etc. The comprehensive method for detecting the microparticles with high concentration is less, the used equipment occupies a large space and is expensive, the equipment cannot be brought to the site for rapid real-time detection, researchers are required to have higher professional knowledge and rich practical experience, the detection speed is slow, and the method has great limitation on the detection of the concentration of the microparticles. The detection of microparticles at high concentrations is therefore the focus of current research.
Disclosure of Invention
In view of the above-mentioned technical problems, a method for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography is provided, which can realize rapid on-site detection of the concentration of the high-concentration microparticles, and has the advantages of simple operation process, low cost and stable identification index.
The technical means adopted by the invention are as follows:
a method for detecting the concentration and activity of high-concentration microparticles based on multi-height lens-free digital holography comprises the following steps:
s1: acquiring original holograms of a plurality of sub-pixel displacements with different z-axis distances in a sample to be detected;
s2: performing sub-pixel registration processing on the obtained original holograms;
s3: based on the original hologram after sub-pixel registration processing, restoring the amplitude and phase information of the image by adopting a single-beam multi-intensity reconstruction algorithm;
s4: performing super-resolution processing on the original hologram by adopting a convex set projection iterative algorithm, and improving the resolution of the hologram;
s5: performing three-dimensional reconstruction on the super-resolution image to obtain the length, width and thickness information of the sample to be detected, and realizing the detection of the activity of the sample to be detected;
s6: and calculating the refractive index of the sample to be detected based on the phase information of the hologram subjected to the super-resolution processing in the step S4 through the relationship between the concentration of each high-concentration microparticle and the refractive index obtained in advance, so as to obtain the concentration of the sample to be detected and realize the detection of the concentration of the high-concentration microparticles.
Further, the step S1 of acquiring a plurality of original holograms with sub-pixel displacement having different z-axis distances includes:
and a plurality of multi-height holographic images with sub-pixel displacement are obtained by controlling the displacement table assembly.
Further, the step S3 specifically includes:
s31, measuring amplitude by using the first measuring plane
Initializing the first measuring plane, i.e.
S32, combining the complex amplitude U of the first measuring plane1Transmitted to a second measuring plane to obtain an estimate of its complex amplitude
The process is described as follows:
U'2(x2,y2)＝((exp(jkd))/(jkd))∫∫U1(x1,y1)exp{((jk)/(2d))[(x2-x1)2+(y2-y1)2]}dx1dy1
s33, keeping the phase unchanged, and replacing the amplitude of the second measuring plane, namely
S34, combining the complex amplitude U of the second measuring plane2Passing to the next plane, and so on until the composite complex amplitude of the Nth measurement plane is generated
S35, measuring the composite complex amplitude U of the Nth measuring planeNThe inverse operation is transmitted to the object plane, so as to obtain a reconstructed light field U of the target0；
S36, reconstructing the complex amplitude U of the target0Transmitting the calculation to the first measurement plane to obtain the complex amplitude estimation value,
and setting a threshold condition and judging by the following formula:
and S37, repeating the steps S31-S36 until the required precision requirement is reached.
Further, the step S4 specifically includes:
s41, selecting the k-th frame in the observed low-resolution image sequence as a reference frame, and interpolating the reference frame to obtain an initial estimation image of the super-resolution image;
s42, performing motion estimation on the low-resolution image sequence and the reference frame to obtain the offset of the low-resolution image relative to the reference frame; calculating the pixel position of each pixel in the low-resolution image, which is mapped to the high-resolution image according to the offset estimation;
s43, according to the PSF function, an estimation value of the current high-resolution image pixel after passing through the imaging system is calculated in a simulating mode, the obtained estimation value is compared with the observed low-resolution pixel value to obtain a residual error, and the high-resolution pixel value is corrected according to the residual error to reduce the residual error to be within an error range;
and S44, iteratively correcting the current high-resolution estimation to obtain the final high-resolution hologram.
Compared with the prior art, the invention has the following advantages:
1. the detection method provided by the invention utilizes the multi-height lens-free digital holographic technology to detect the concentration and activity of high-concentration microparticles, combines single-beam multi-intensity reconstruction with a convex set projection iterative algorithm, and compared with other holographic three-dimensional morphology technologies, the method performs super-resolution reconstruction during image processing, so that the three-dimensional morphology is recovered more accurately, errors of final measurement results are reduced, and the method does not need a unwrapping step, greatly simplifies data processing steps, and saves time.
2. The method for detecting the concentration and the activity of the high-concentration microparticles based on the multi-height lens-free digital holography can obtain the two-dimensional and three-dimensional shapes of the samples. But compared with an optical microscope, the system is suitable for on-site rapid detection and has the advantages of low price, light weight and small volume.
3. The method for detecting the concentration and the activity of the high-concentration microparticles based on the multi-height lens-free digital holography can obtain the three-dimensional morphology of a sample, can detect the activity of the high-concentration microparticles according to the characteristics and relevant information of the three-dimensional morphology, and is simple to operate and wide in application range.
Based on the reasons, the invention can be widely popularized in the fields of high-concentration microparticle concentration and activity detection and the like.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed to be used in the description of the embodiments or the prior art will be briefly introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to these drawings without creative efforts.
Fig. 1 is a schematic structural diagram of a high-concentration microparticle concentration and activity detection device based on multi-height lensless digital holography according to an embodiment of the present invention.
Fig. 2 is a schematic structural diagram of a microfluidic chip in the detection device provided in the embodiment of the present invention.
FIG. 3 is a flow chart of the method of the present invention.
Fig. 4 is a flowchart of steps of a single-beam multi-intensity reconstruction algorithm according to an embodiment of the present invention.
In the figure: 1. a light source driving assembly; 2. a light source assembly; 3. a microporous component; 4. a light propagating assembly; 5. a sample stage assembly; 6. a displacement table assembly; 7. an image acquisition component 8 and an image processing component; 9. polydimethylsiloxane; 10. a glass slide; 11. a sample tank; 12. detecting a region; 13. a waste liquid tank.
Detailed Description
In order to make the technical solutions of the present invention better understood, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
It should be noted that the terms "first," "second," and the like in the description and claims of the present invention and in the drawings described above are used for distinguishing between similar elements and not necessarily for describing a particular sequential or chronological order. It is to be understood that the data so used is interchangeable under appropriate circumstances such that the embodiments of the invention described herein are capable of operation in sequences other than those illustrated or described herein. Furthermore, the terms "comprises," "comprising," and "having," and any variations thereof, are intended to cover a non-exclusive inclusion, such that a process, method, system, article, or apparatus that comprises a list of steps or elements is not necessarily limited to those steps or elements expressly listed, but may include other steps or elements not expressly listed or inherent to such process, method, article, or apparatus.
The invention relates to a method and a device for detecting the concentration and activity of high-concentration microparticles based on multi-height lens-free digital holography, which are based on the principle that holographic images formed by high-concentration microparticles with different concentrations have different holographic pattern characteristics and can obtain holographic images with higher resolution by processing a plurality of multi-height holographic images, and is further described with reference to the attached drawings.
As shown in fig. 1, the structure of the high-concentration microparticle concentration and activity detection device based on multi-height lensless digital holography is schematically illustrated, and the device comprises a light source driving component 1, a light source 2, a micropore component 3, a light propagation component 4, a sample stage component 5, a displacement stage component 6, an image acquisition component 7 and an image processing component 8 which are connected in sequence; the light source driving assembly 1 excites the light source 2 to emit blue light, the light penetrates through the micropore assembly 3 to disperse partial coherent light into spherical waves, the spherical waves are transmitted for a distance through the light transmission assembly 4 and then reach the sample stage assembly 5, the distance between a sample and the image acquisition assembly 7 is controlled through the displacement stage assembly 6, and then holographic images with different z-axis heights are formed on the image acquisition assembly 7; (reference light waves do not need to be independently introduced, the transmitted light of the object is the reference light waves, the scattered light is the object light waves, and the reference light waves and the object light waves are mutually interfered to form a holographic image in the image acquisition assembly 7). the image processing assembly 8 carries out three-dimensional shape recovery and characteristic analysis on the obtained holographic pattern so as to detect the concentration and activity of the high-concentration microparticles. The amplitude and phase information of the image are restored through a single-beam multi-intensity reconstruction (SBMIR) algorithm, the holographic image is subjected to super-resolution processing through a convex set projection iterative algorithm, the resolution of the holographic image can be improved, the three-dimensional appearance of the sample can be finally restored, and the length, width and thickness information of the sample can be obtained through the three-dimensional appearance.
In specific implementation, as a preferred embodiment of the present invention, as shown in fig. 2, the microfluidic chip in the sample stage assembly includes a polydimethylsiloxane sheet 9 and a glass slide 10, the polydimethylsiloxane sheet 9 is sequentially engraved with a detection region 11, and two ends of the detection region are symmetrically connected to a sample tank 12 and a waste liquid tank 13. The use of the microfluidic chip can ensure that the sample is not seriously layered and the sample is prevented from being polluted.
As shown in fig. 3, the present invention provides a method for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography, which comprises the following steps:
s1: acquiring original holograms of a plurality of sub-pixel displacements with different z-axis distances in a sample to be detected;
s2: performing sub-pixel registration processing on the obtained original holograms;
s3: based on the original hologram after sub-pixel registration processing, restoring the amplitude and phase information of the image by adopting a single-beam multi-intensity reconstruction algorithm;
in specific implementation, as a preferred embodiment of the present invention, as shown in fig. 4, the step S3 specifically includes:
s31, measuring amplitude by using the first measuring plane
Initializing the first measuring plane, i.e.
S32, combining the complex amplitude U of the first measuring plane1Is transmitted toSecond measuring plane to obtain the estimated value of its complex amplitude
The process is described as follows:
U'2(x2,y2)＝((exp(jkd))/(jkd))∫∫U1(x1,y1)exp{((jk)/(2d))[(x2-x1)2+(y2-y1)2]}dx1dy1
s33, keeping the phase unchanged, and replacing the amplitude of the second measuring plane, namely
S34, combining the complex amplitude U of the second measuring plane2Passing to the next plane, and so on until the composite complex amplitude of the Nth measurement plane is generated
S35, measuring the composite complex amplitude U of the Nth measuring planeNThe inverse operation is transmitted to the object plane, so as to obtain a reconstructed light field U of the target0；
S36, reconstructing the complex amplitude U of the target0Transmitting the calculation to the first measurement plane to obtain the complex amplitude estimation value,
and setting a threshold condition and judging by the following formula:
and S37, repeating the steps S31-S36 until the required precision requirement is reached.
S4: performing super-resolution processing on the original hologram by adopting a convex set projection iterative algorithm, and improving the resolution of the hologram;
in a specific implementation, as a preferred embodiment of the present invention, the step S4 specifically includes:
s41, selecting the k-th frame in the observed low-resolution image sequence as a reference frame, and interpolating the reference frame to obtain an initial estimation image of the super-resolution image;
s42, performing motion estimation on the low-resolution image sequence and the reference frame to obtain the offset of the low-resolution image relative to the reference frame; calculating the pixel position of each pixel in the low-resolution image, which is mapped to the high-resolution image according to the offset estimation;
s43, according to the PSF function, an estimation value of the current high-resolution image pixel after passing through the imaging system is calculated in a simulating mode, the obtained estimation value is compared with the observed low-resolution pixel value to obtain a residual error, and the high-resolution pixel value is corrected according to the residual error to reduce the residual error to be within an error range;
and S44, iteratively correcting the current high-resolution estimation to obtain the final high-resolution hologram.
S5: performing three-dimensional reconstruction on the super-resolution image to obtain the length, width and thickness information of the sample to be detected, and realizing the detection of the activity of the sample to be detected;
s6: and calculating the refractive index of the sample to be detected based on the phase information of the hologram subjected to the super-resolution processing in the step S4 through the relationship between the concentration of each high-concentration microparticle and the refractive index obtained in advance, so as to obtain the concentration of the sample to be detected and realize the detection of the concentration of the high-concentration microparticles.
Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit the same; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.
Claims (6)
1. A high-concentration microparticle concentration and activity detection method based on multi-height lens-free digital holography is characterized by comprising the following steps:
s1: acquiring original holograms of a plurality of sub-pixel displacements with different z-axis distances in a sample to be detected;
s2: performing sub-pixel registration processing on the obtained original holograms;
s3: based on the original hologram after sub-pixel registration processing, restoring the amplitude and phase information of the image by adopting a single-beam multi-intensity reconstruction algorithm;
s4: performing super-resolution processing on the original hologram by adopting a convex set projection iterative algorithm, and improving the resolution of the hologram;
s5: performing three-dimensional reconstruction on the super-resolution image to obtain the length, width and thickness information of the sample to be detected, and realizing the detection of the activity of the sample to be detected;
s6: and calculating the refractive index of the sample to be detected based on the phase information of the hologram subjected to the super-resolution processing in the step S4 through the relationship between the concentration of each high-concentration microparticle and the refractive index obtained in advance, so as to obtain the concentration of the sample to be detected and realize the detection of the concentration of the high-concentration microparticles.
2. The method for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography according to claim 1, wherein the step S1 of obtaining a plurality of original holograms with different z-axis distances and sub-pixel displacements comprises:
and a plurality of multi-height holographic images with sub-pixel displacement are obtained by controlling the displacement table assembly.
3. The method for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography according to claim 1, wherein the step S3 specifically comprises:
s31, measuring amplitude by using the first measuring plane
Initializing the first measuring plane, i.e.
S32, combining the complex amplitude U of the first measuring plane1Transmitted to a second measuring plane to obtain an estimate of its complex amplitude
The process is described as follows:
U′2(x2,y2)＝((exp(jkd))/(jkd))∫∫U1(x1,y1)exp{((jk)/(2d))[(x2-x1)2+(y2-y1)2]}dx1dy1
s33, keeping the phase unchanged, and replacing the amplitude of the second measuring plane, namely
S34, combining the complex amplitude U of the second measuring plane2Passing to the next plane, and so on until the composite complex amplitude of the Nth measurement plane is generated
S35, measuring the composite complex amplitude U of the Nth measuring planeNThe inverse operation is transmitted to the object plane, so as to obtain a reconstructed light field U of the target0；
S36, reconstructing the complex amplitude U of the target0Transmitting the calculation to the first measurement plane to obtain the complex amplitude estimation value,
and setting a threshold condition and judging by the following formula:
and S37, repeating the steps S31-S36 until the required precision requirement is reached.
4. The method for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography according to claim 1, wherein the step S4 specifically comprises:
s41, selecting the k-th frame in the observed low-resolution image sequence as a reference frame, and interpolating the reference frame to obtain an initial estimation image of the super-resolution image;
s42, performing motion estimation on the low-resolution image sequence and the reference frame to obtain the offset of the low-resolution image relative to the reference frame; calculating the pixel position of each pixel in the low-resolution image, which is mapped to the high-resolution image according to the offset estimation;
s43, according to the PSF function, an estimation value of the current high-resolution image pixel after passing through the imaging system is calculated in a simulating mode, the obtained estimation value is compared with the observed low-resolution pixel value to obtain a residual error, and the high-resolution pixel value is corrected according to the residual error to reduce the residual error to be within an error range;
and S44, iteratively correcting the current high-resolution estimation to obtain the final high-resolution hologram.
5. The device for detecting the concentration and activity of the high-concentration microparticles based on the multi-height lens-free digital holography is characterized in that the method for detecting the concentration and activity of the high-concentration microparticles based on the multi-height lens-free digital holography, which is disclosed by any one of claims 1 to 4, is repeatedly executed, and comprises the following steps: the device comprises a light source driving component (1), a light source (2), a micropore component (3), a light propagation component (4), a sample stage component (5), a displacement stage component (6), an image acquisition component (7) and an image processing component (8) which are connected in sequence;
the light source driving assembly (1) is characterized in that an excitation light source (2) emits blue light, the light penetrates through the micropore assembly (3) to disperse partial coherent light into spherical waves, the spherical waves are transmitted for a distance through the light transmission assembly (4) and then reach the sample table assembly (5), the distance between a sample and the image acquisition assembly (7) is controlled through the displacement table assembly (6), and holographic images with different z-axis heights are formed in the image acquisition assembly (7); and the image processing component (8) is used for carrying out three-dimensional shape recovery and characteristic analysis on the obtained holographic pattern so as to detect the concentration and activity of the high-concentration microparticles.
6. The device for detecting the concentration and activity of high-concentration microparticles based on multi-height lensless digital holography according to claim 5, wherein the sample stage assembly comprises a microfluidic chip, the microfluidic chip comprises a polydimethylsiloxane sheet (9) and a glass slide (10), the polydimethylsiloxane sheet (9) is sequentially notched with a detection area (11), and two ends of the detection area are symmetrically connected with the sample groove (12) and the waste liquid groove (13).
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113311682A (en) * | 2021-05-08 | 2021-08-27 | 江苏大学 | Lens-free coaxial digital holographic microparticle imaging system and method |
| CN115984341A (en) * | 2023-03-20 | 2023-04-18 | 深圳市朗诚科技股份有限公司 | Marine water quality microorganism detection method, device, equipment and storage medium |
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| CN108983579A (en) * | 2018-09-05 | 2018-12-11 | 南京大学 | Method and device thereof without lens digital holographic microscopic imaging phase recovery and reconstruction |
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| CN115984341A (en) * | 2023-03-20 | 2023-04-18 | 深圳市朗诚科技股份有限公司 | Marine water quality microorganism detection method, device, equipment and storage medium |
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