CN113804625A - Automatic tracking cell imaging method and system - Google Patents
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Abstract
The invention belongs to the technical field of vortex rotation imaging, and particularly relates to an automatic tracking cell imaging method and system. The automatic tracking cell imaging method comprises the following steps: performing edge-enhanced imaging on the cells; collecting cell images; obtaining cell change information; and adjusting the edge enhancement direction according to the cell change information. The invention has the advantages that the invention realizes the anisotropic edge enhancement by using the spatial light modulator to load the Bessel-like superposition vortex filter on the frequency spectrum surface of the imaging system; and meanwhile, images received by the CCD are observed, when the cells change, the change information is extracted and processed, and the processed result is reloaded on the spatial light modulator, so that the edge enhancement direction of the edge enhancement module is consistent with the edge enhancement direction required by the cells, and the automatic tracking imaging of the cell change can be realized.
Description
Technical Field
The invention belongs to the technical field of vortex rotation imaging, and particularly relates to an automatic tracking cell imaging method and system.
Background
In scientific research, we often need to image some tiny things, including amplitude type objects and phase type objects. The traditional imaging technology has weak processing capability on most phase type biological samples, and because the light beam only changes the phase after penetrating through a phase type object and the amplitude does not change obviously, human eyes cannot distinguish the physical structure of the substance to be detected. Imaging often requires direct contact with the sample, which can contaminate the sample, or requires observation using a complex designed system, which is more costly.
By adopting two-dimensional radial Hilbert transform, the optical system can be very sensitive to the phase gradient change of a detected sample, and the spatial distribution of the phase gradient can be converted into an image with intensity change, so that the imaging quality of the sample is greatly improved. The spiral phase plate expands the one-dimensional Hilbert transform to a radial space, and an isotropic edge enhancement effect can be obtained by performing vortex filtering on an image in a spatial frequency spectrum plane. If features in a certain direction need to be emphasized, an anisotropic edge enhancement technique, anisotropic, is required.
Many anisotropic edge enhancement techniques are proposed, such as fractional order vortex filters, off-axis vortex filters, etc., which achieve edge enhancement in one direction. However, in actual imaging, when the cell is a living cell and changes from moment to moment, the above-described anisotropic edge enhancement is used, and manual adjustment is necessary after the cell change, which is very inflexible.
Disclosure of Invention
The invention aims to provide an automatic tracking cell imaging method and system.
In order to solve the above technical problem, the present invention provides an automatic tracking cell imaging method, comprising: performing edge-enhanced imaging on the cells; collecting cell images; obtaining cell change information; and adjusting the edge enhancement direction according to the cell change information.
In yet another aspect, the present invention also provides an automatic tracking cell imaging system, comprising: a processor module; a cell imaging module for imaging a cell; the edge enhancement module is used for carrying out edge enhancement on cell imaging; the imaging acquisition module is used for acquiring cell images after edge enhancement and transmitting the cell images to the processor module; the processor module is adapted to adjust an edge enhancement direction of the edge enhancement module according to a desired edge enhancement direction of the cell.
The invention has the advantages that the invention realizes the anisotropic edge enhancement by using the spatial light modulator to load the Bessel-like superposition vortex filter on the frequency spectrum surface of the imaging system; and meanwhile, images received by the CCD are observed, when the cells change, the change information is extracted and processed, and the processed result is reloaded on the spatial light modulator, so that the edge enhancement direction of the edge enhancement module is consistent with the edge enhancement direction required by the cells, and the automatic tracking imaging of the cell change can be realized.
Additional features and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objectives and other advantages of the invention will be realized and attained by the structure particularly pointed out in the written description and claims hereof as well as the appended drawings.
In order to make the aforementioned and other objects, features and advantages of the present invention comprehensible, preferred embodiments accompanied with figures are described in detail below.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a flow chart of a method of automated follow-up cell imaging in accordance with the present invention;
FIG. 2 is a schematic diagram of a cellular imaging optical path in accordance with the present invention;
FIG. 3 is an optical schematic of cellular imaging according to the present invention;
FIG. 4 is a computed hologram of a Bessel-like superimposed vortex filter according to the present invention;
FIG. 5 is a schematic diagram of a cellular transformation process according to the present invention;
FIG. 6 is a graph of point spread function simulations of Bessel-like superimposed vortices in accordance with the present invention;
FIG. 7 is a flow chart of a preferred embodiment of an automated tracking cell imaging method in accordance with the present invention;
FIG. 8 is a schematic diagram of the direction of edge enhancement of a Bessel-like superposition vortex filter and the direction of cell-required edge enhancement according to the present invention;
FIG. 9 is a graph showing the effect of dynamic tracking imaging when cells are changed according to the present invention;
FIG. 10 is a schematic diagram of an automated tracking cell imaging system in accordance with the present invention.
Detailed Description
To make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings, and it is apparent that the described embodiments are some, but not all embodiments of the present invention. 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.
As shown in fig. 1, in one embodiment, the present invention provides an automated tracking cell imaging method, comprising: performing edge-enhanced imaging on the cells; collecting cell images; obtaining cell change information; and adjusting the edge enhancement direction according to the cell change information.
As shown in fig. 2, fig. 3 and fig. 4, in this embodiment, preferably, the performing edge enhanced imaging on the cell includes: the system comprises a laser, an attenuator, a beam expander, an aperture diaphragm, two lenses with the same focal length and a spatial light modulator positioned on a confocal frequency spectrum plane position of the two lenses, wherein the laser, the attenuator, the beam expander, the aperture diaphragm, the two lenses with the same focal length and the spatial light modulator are sequentially arranged; loading a computed hologram of a Bessel-like superposition vortex filter on the spatial light modulator; the cell sample is placed at the focal length of the first lens in the imaging optical path, and the filtered cell image is output at the back focal length of the second lens.
In this embodiment, anisotropic edge enhancement is achieved by loading a bessel-like superposition vortex filter at the confocal spectral plane position of the two lenses using a spatial light modulator.
In this embodiment, the computed hologram is generated from a representation of the transmittance function of a vortex filter of a bessel-like superimposed vortex filter; the expression of the transmittance function includes:
wherein:
(ρ, φ) is a polar coordinate system of the spectral plane;
J2(α ρ) is a second order Bessel function of the first kind;
alpha is a modulation parameter for adjusting J2The value of (α ρ) ranges from 0 to 10000, which can be 300 in this embodiment;
exp[il(φ+φ0)]and exp [ -il (φ + φ)0)]Is the phase factor of the vortex; one of which may be a positive vortex and the other a negative vortex;
c is a weight coefficient of the weight ratio of the positive vortex and the negative vortex, the value range is 0-1, when c is 1, the situation is symmetrical, when c is 0, the isotropic edge enhancement is realized, and in the embodiment, 1 can be obtained; i represents an imaginary number; l is the topological charge number of the vortex phase, the value range is a positive integer, and can be 1, 2, 3 or 60, 64, etc., and can be 1 in this embodiment; phi is a0Representing an initial azimuth;
as shown in fig. 2, the cell image may be collected but not limited to be magnified by two objective lenses, and the magnification factor may be selected as required; the amplified optical signal for cell imaging is then converted into an analog current signal by a CCD charge-coupled device and transmitted to a processor module, which may be a computer.
As shown in fig. 5, in actual imaging, when the cell is a living cell and changes all the time, the cell change needs to be tracked, the edge enhancement direction of the bessel-like superposition vortex filter needs to be adjusted in time, and the edge enhancement direction of the bessel-like superposition vortex filter is adjusted to be consistent with the edge enhancement direction of the cell, so that the cell imaging can be kept clear in real time.
In this embodiment, the acquiring cell change information includes: in the first imaging, the edge enhancement direction of the Bessel-like superposition vortex filter and the edge enhancement direction required by the cell can be adjusted to be consistent manually, and the edge enhancement direction required by the cell is marked; obtaining the cell after cell movement requires a deflection angle of the edge enhancement direction compared to the edge enhancement direction of the bessel-like superposition vortex filter.
In this embodiment, the edge enhancement direction of the bessel-like superposition vortex filter is obtained by:
fourier transformation is carried out on the transmittance function of the Bessel-like superposed vortex filter to obtain a point spread function of the Bessel-like superposed vortex filter, and the expression of the point spread function is as follows:
wherein: λ is the wavelength of the laser, f is the focal length of the lens,
is a first order Bessel function of the first kind;
and (5) simulating a point spread function, wherein the directions of the two light spots are the edge enhancement directions of the vortex filter. As shown in fig. 6. In an optical system, the light field distribution of an output image of a point light source as an input object is referred to as a point spread function.
In this embodiment, the adjusting the edge enhancement direction according to the cell change information includes: adjusting phi in the expression of the transmittance function according to the deflection angle0To adjust the edge enhancement direction of the bessel-like superposition vortex filter to be consistent with the required edge enhancement direction of the cell after the cell movement.
In a preferred embodiment, as shown in fig. 7, in the first imaging, the edge enhancement direction of the bessel-like superposition vortex filter can be adjusted to be consistent with the edge enhancement direction required by the cell by manual operation, and the result is marked by the computer terminal.
As shown in the (1) th sub-diagram in fig. 8, two marker points may be selected on the primary CCD imaging result, the two marker points are connected to form a straight line, the direction of the straight line is determined as the cell required edge enhancement direction, the edge enhancement direction of the bessel-like superimposing vortex filter is indicated by a dotted line, and the edge enhancement direction is adjusted to be consistent with the cell required edge enhancement direction.
When the cell changes, the positions of the two mark points are tracked, the two mark points are connected into a straight line, and the direction in which the two mark points are connected into the straight line is recorded, namely the change of the solid line in fig. 8 is the real-time change angle and direction of the cell in the direction needing edge enhancement.
And comparing the solid line with the dotted line at the computer terminal to obtain the angle and the direction between the solid line and the dotted line. When the dotted line and the thin solid line completely coincide, the edge enhancement effect is the best. When the cell sample changes and the two directions are not consistent, the imaging is unclear, and the edge enhancement direction (dotted line) of the Bessel-like superposition vortex filter and the edge enhancement direction (solid line) required by the cell need to be adjusted to be consistent.
Taking the sub-diagram (2) in FIG. 8 as an example, the direction (solid line) in which the cell needs to be edge-enhanced obtained by the computer terminal is compared with the direction (dotted line) in which the edge of the Bezier-like superimposed vortex filter is enhancedThe offset direction is counterclockwise and the offset angle is 10 °. From the expression of the transmittance function of the Bessel-like superposition vortex filter, phi0Indicating the initial azimuth by varying phi0The Bessel-like superposition vortex filter edge enhancement direction can be changed. In this example, phi in the expression of the Bessel-like superposition vortex filter can be changed0A value of (d), will0Becomes phi0And +10 degrees (the counter-clockwise direction can be defined as addition, and the clockwise direction can be defined as subtraction), then the computer hologram is regenerated at the computer terminal, and then the newly generated computer hologram is loaded on the spatial light modulator, so that the edge enhancement direction of the Bessel-like superposition vortex filter can be completely consistent with the edge enhancement direction required by the cell. The adjustment of the sub-diagrams (3) and (4) in fig. 8 is the same as the adjustment of the sub-diagram (2).
In this embodiment, an image received by the CCD is observed, and when a cell changes, change information is extracted and processed, and the processed result is reloaded onto the spatial light modulator, so that the edge enhancement direction of the edge enhancement module is consistent with the direction in which the cell needs edge enhancement, and automatic tracking imaging of the cell change can be realized.
In the embodiment, anisotropic edge enhancement can be realized, namely, the edge in a certain direction is enhanced, and higher image edge contrast and lower diffraction noise are realized; the orientation can also be adjusted by following the cell activity.
On the basis of the above embodiment, the present invention further provides an automatic tracking cell imaging system, including: a processor module; a cell imaging module for imaging a cell; the edge enhancement module is used for carrying out edge enhancement on cell imaging; the imaging acquisition module is used for acquiring cell images after edge enhancement and transmitting the cell images to the processor module; the processor module is adapted to adjust an edge enhancement direction of the edge enhancement module according to a desired edge enhancement direction of the cell.
In this embodiment, the cell imaging system can image the cell by using the above-mentioned cell imaging method.
In this embodiment, optionally, the cell imaging module may include: the system comprises a laser, an attenuator, a beam expander, an aperture diaphragm, two lenses with the same focal length and a spatial light modulator positioned on a frequency spectrum plane position confocal by the two lenses, which are sequentially arranged.
In this embodiment, optionally, the edge enhancement module includes a bessel superposition vortex filter electrically connected to the processor module, and is configured to load the spatial light modulator with the computation hologram.
In this embodiment, optionally, the imaging acquisition module includes: an objective lens and a CCD; the CCD is used for collecting cell images amplified by the objective lens and transmitting the cell images to the processor module.
In this embodiment, optionally, a beam splitter may be further disposed between the two lenses with equal focal lengths to divert the light, so as to facilitate the optical path transmission between the modules.
In conclusion, the invention realizes anisotropic edge enhancement by using the spatial light modulator to load the Bessel-like superposition vortex filter on the frequency spectrum surface of the imaging system; and meanwhile, images received by the CCD are observed, when the cells change, the change information is extracted and processed, and the processed result is reloaded on the spatial light modulator, so that the edge enhancement direction of the edge enhancement module is consistent with the edge enhancement direction required by the cells, and the automatic tracking imaging of the cell change can be realized.
In the embodiments provided in the present application, it should be understood that the disclosed apparatus and method can be implemented in other ways. The apparatus embodiments described above are merely illustrative, and for example, the flowchart and block diagrams in the figures illustrate the architecture, functionality, and operation of possible implementations of apparatus, methods and computer program products according to various embodiments of the present invention. In this regard, each block in the flowchart or block diagrams may represent a module, segment, or portion of code, which comprises one or more executable instructions for implementing the specified logical function(s). It should also be noted that, in some alternative implementations, the functions noted in the block may occur out of the order noted in the figures. For example, two blocks shown in succession may, in fact, be executed substantially concurrently, or the blocks may sometimes be executed in the reverse order, depending upon the functionality involved. It will also be noted that each block of the block diagrams and/or flowchart illustration, and combinations of blocks in the block diagrams and/or flowchart illustration, can be implemented by special purpose hardware-based systems which perform the specified functions or acts, or combinations of special purpose hardware and computer instructions.
In addition, the functional modules in the embodiments of the present invention may be integrated together to form an independent part, or each module may exist separately, or two or more modules may be integrated to form an independent part.
The functions, if implemented in the form of software functional modules and sold or used as a stand-alone product, may be stored in a computer readable storage medium. Based on such understanding, the technical solution of the present invention may be embodied in the form of a software product, which is stored in a storage medium and includes instructions for causing a computer device (which may be a personal computer, a server, or a network device) to execute all or part of the steps of the method according to the embodiments of the present invention. And the aforementioned storage medium includes: a U-disk, a removable hard disk, a Read-Only Memory (ROM), a Random Access Memory (RAM), a magnetic disk or an optical disk, and other various media capable of storing program codes.
In light of the foregoing description of the preferred embodiment of the present invention, many modifications and variations will be apparent to those skilled in the art without departing from the spirit and scope of the invention. The technical scope of the present invention is not limited to the content of the specification, and must be determined according to the scope of the claims.
Claims (10)
1. An automated tracking cell imaging method, comprising:
performing edge-enhanced imaging on the cells;
collecting cell images;
obtaining cell change information;
and adjusting the edge enhancement direction according to the cell change information.
2. The method of automated tracked cellular imaging according to claim 1,
the edge-enhanced imaging of cells comprises:
the system comprises a laser, an attenuator, a beam expander, an aperture diaphragm, two lenses with the same focal length and a spatial light modulator positioned on a confocal frequency spectrum plane position of the two lenses, wherein the laser, the attenuator, the beam expander, the aperture diaphragm, the two lenses with the same focal length and the spatial light modulator are sequentially arranged;
loading a computed hologram of a Bessel-like superposition vortex filter on the spatial light modulator;
the cell sample is placed at the focal length of the first lens in the imaging optical path, and the filtered cell image is output at the back focal length of the second lens.
3. The method of automated tracked cellular imaging according to claim 2,
the calculation hologram is generated according to a transmittance function expression of the vortex filter of the Bezier-like superposition vortex filter;
the expression of the transmittance function includes:
wherein:
(ρ, φ) is a polar coordinate system of the spectral plane;
J2(α ρ) is a second order Bessel function of the first kind;
alpha is a modulation parameter for adjusting J2The value of (alpha rho) ranges from 0 to 10000;
exp[il(φ+φ0)]and exp [ -il (φ + φ)0)]Is swirledA phase factor;
c is a weight coefficient of the weight ratio of the positive vortex and the negative vortex, the value range is 0-1, when c is 1, the situation is symmetrical, and when c is 0, the isotropic edge enhancement is realized; i represents an imaginary number; l is the topological charge number of the vortex phase, and the value range is a positive integer; phi is a0Representing an initial azimuth;
4. the method of automated tracked cellular imaging according to claim 3,
the acquiring cell change information includes:
in the first imaging, the edge enhancement direction of the Bessel-like superposition vortex filter is adjusted to be consistent with the edge enhancement direction required by the cell, and the edge enhancement direction required by the cell is marked;
obtaining the cell after cell movement requires a deflection angle of the edge enhancement direction compared to the edge enhancement direction of the bessel-like superposition vortex filter.
5. The method of automated tracked cellular imaging according to claim 4,
the edge enhancement direction of the Bessel-like superposition vortex filter is obtained by the following steps:
fourier transformation is carried out on the transmittance function of the Bessel-like superposed vortex filter to obtain a point spread function of the Bessel-like superposed vortex filter, and the expression of the point spread function is as follows:
wherein: (r, θ) is the polar coordinate system of the cell imaging plane; λ is the wavelength of the laser, f is the focal length of the lens,
is a first order Bessel function of the first kind;
and (5) simulating a point spread function, wherein the directions of the two light spots are the edge enhancement directions of the vortex filter.
6. The method of automated tracked cellular imaging according to claim 5,
the adjusting the edge enhancement direction according to the cell change information comprises:
adjusting phi in the expression of the transmittance function according to the deflection angle0To adjust the edge enhancement direction of the bessel-like superposition vortex filter to be consistent with the required edge enhancement direction of the cell after the cell movement.
7. An automated tracking cell imaging system, comprising:
a processor module;
a cell imaging module for imaging a cell;
the edge enhancement module is used for carrying out edge enhancement on cell imaging;
the imaging acquisition module is used for acquiring cell images after edge enhancement and transmitting the cell images to the processor module;
the processor module is adapted to adjust an edge enhancement direction of the edge enhancement module according to a desired edge enhancement direction of the cell.
8. The automated tracked cell imaging system of claim 7,
the cell imaging module includes: the system comprises a laser, an attenuator, a beam expander, an aperture diaphragm, two lenses with the same focal length and a spatial light modulator positioned on a frequency spectrum plane position confocal by the two lenses, which are sequentially arranged.
9. The automated tracked cell imaging system of claim 8,
the edge enhancement module comprises a Bezier superposition vortex filter electrically connected with the processor module and is used for loading the spatial light modulator with a calculation hologram.
10. The automated tracked cell imaging system of claim 9,
the imaging acquisition module comprises: an objective lens and a CCD;
the CCD is used for collecting cell images amplified by the objective lens and transmitting the cell images to the processor module.
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