CN111122446A - Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity - Google Patents
Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity Download PDFInfo
- Publication number
- CN111122446A CN111122446A CN201911089768.XA CN201911089768A CN111122446A CN 111122446 A CN111122446 A CN 111122446A CN 201911089768 A CN201911089768 A CN 201911089768A CN 111122446 A CN111122446 A CN 111122446A
- Authority
- CN
- China
- Prior art keywords
- cavity
- detected
- cell
- absorption rate
- path
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000010521 absorption reaction Methods 0.000 title claims abstract description 75
- 238000010998 test method Methods 0.000 title claims abstract description 22
- 230000003287 optical effect Effects 0.000 claims abstract description 30
- 238000000034 method Methods 0.000 claims abstract description 25
- 238000005259 measurement Methods 0.000 claims abstract description 14
- 238000012545 processing Methods 0.000 claims abstract description 5
- 238000004364 calculation method Methods 0.000 claims description 9
- 238000002310 reflectometry Methods 0.000 claims description 5
- 230000009466 transformation Effects 0.000 claims description 5
- 239000013307 optical fiber Substances 0.000 claims description 3
- 238000009795 derivation Methods 0.000 claims description 2
- 230000001413 cellular effect Effects 0.000 claims 2
- 238000000691 measurement method Methods 0.000 claims 1
- 244000005700 microbiome Species 0.000 claims 1
- 238000003384 imaging method Methods 0.000 abstract description 22
- 230000035945 sensitivity Effects 0.000 abstract description 12
- 238000012360 testing method Methods 0.000 abstract description 9
- 238000003325 tomography Methods 0.000 abstract description 9
- BJQHLKABXJIVAM-UHFFFAOYSA-N bis(2-ethylhexyl) phthalate Chemical compound CCCCC(CC)COC(=O)C1=CC=CC=C1C(=O)OCC(CC)CCCC BJQHLKABXJIVAM-UHFFFAOYSA-N 0.000 description 14
- 238000010586 diagram Methods 0.000 description 7
- 238000005516 engineering process Methods 0.000 description 7
- 241000223785 Paramecium Species 0.000 description 4
- 238000012576 optical tweezer Methods 0.000 description 4
- 238000011160 research Methods 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 2
- 239000005357 flat glass Substances 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 238000001093 holography Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000004075 alteration Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 238000002591 computed tomography Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000000799 fluorescence microscopy Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012014 optical coherence tomography Methods 0.000 description 1
- 230000000149 penetrating effect Effects 0.000 description 1
- 230000010363 phase shift Effects 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
-
- G—PHYSICS
- G03—PHOTOGRAPHY; CINEMATOGRAPHY; ANALOGOUS TECHNIQUES USING WAVES OTHER THAN OPTICAL WAVES; ELECTROGRAPHY; HOLOGRAPHY
- G03H—HOLOGRAPHIC PROCESSES OR APPARATUS
- G03H1/00—Holographic processes or apparatus using light, infrared or ultraviolet waves for obtaining holograms or for obtaining an image from them; Details peculiar thereto
- G03H1/0005—Adaptation of holography to specific applications
-
- G—PHYSICS
- G03—PHOTOGRAPHY; CINEMATOGRAPHY; ANALOGOUS TECHNIQUES USING WAVES OTHER THAN OPTICAL WAVES; ELECTROGRAPHY; HOLOGRAPHY
- G03H—HOLOGRAPHIC PROCESSES OR APPARATUS
- G03H1/00—Holographic processes or apparatus using light, infrared or ultraviolet waves for obtaining holograms or for obtaining an image from them; Details peculiar thereto
- G03H1/04—Processes or apparatus for producing holograms
- G03H1/0443—Digital holography, i.e. recording holograms with digital recording means
-
- G—PHYSICS
- G03—PHOTOGRAPHY; CINEMATOGRAPHY; ANALOGOUS TECHNIQUES USING WAVES OTHER THAN OPTICAL WAVES; ELECTROGRAPHY; HOLOGRAPHY
- G03H—HOLOGRAPHIC PROCESSES OR APPARATUS
- G03H1/00—Holographic processes or apparatus using light, infrared or ultraviolet waves for obtaining holograms or for obtaining an image from them; Details peculiar thereto
- G03H1/22—Processes or apparatus for obtaining an optical image from holograms
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N2021/1765—Method using an image detector and processing of image signal
- G01N2021/177—Detector of the video camera type
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N2021/178—Methods for obtaining spatial resolution of the property being measured
- G01N2021/1785—Three dimensional
- G01N2021/1787—Tomographic, i.e. computerised reconstruction from projective measurements
-
- G—PHYSICS
- G03—PHOTOGRAPHY; CINEMATOGRAPHY; ANALOGOUS TECHNIQUES USING WAVES OTHER THAN OPTICAL WAVES; ELECTROGRAPHY; HOLOGRAPHY
- G03H—HOLOGRAPHIC PROCESSES OR APPARATUS
- G03H1/00—Holographic processes or apparatus using light, infrared or ultraviolet waves for obtaining holograms or for obtaining an image from them; Details peculiar thereto
- G03H1/0005—Adaptation of holography to specific applications
- G03H2001/0033—Adaptation of holography to specific applications in hologrammetry for measuring or analysing
- G03H2001/0038—Adaptation of holography to specific applications in hologrammetry for measuring or analysing analogue or digital holobjects
-
- G—PHYSICS
- G03—PHOTOGRAPHY; CINEMATOGRAPHY; ANALOGOUS TECHNIQUES USING WAVES OTHER THAN OPTICAL WAVES; ELECTROGRAPHY; HOLOGRAPHY
- G03H—HOLOGRAPHIC PROCESSES OR APPARATUS
- G03H1/00—Holographic processes or apparatus using light, infrared or ultraviolet waves for obtaining holograms or for obtaining an image from them; Details peculiar thereto
- G03H1/0005—Adaptation of holography to specific applications
- G03H2001/005—Adaptation of holography to specific applications in microscopy, e.g. digital holographic microscope [DHM]
Landscapes
- Physics & Mathematics (AREA)
- General Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Engineering & Computer Science (AREA)
- Computing Systems (AREA)
- Theoretical Computer Science (AREA)
- Investigating Or Analysing Materials By Optical Means (AREA)
Abstract
The invention provides a novel three-dimensional test method for the common-path F-P cavity enhanced cell absorption rate. The method is characterized in that: the method comprises the steps of digital hologram recording, numerical reconstruction, error processing and three-dimensional absorptivity distribution reconstruction based on an F-P cavity. The invention mainly provides a novel method for three-dimensional testing of the common-path F-P cavity enhanced cell absorption rate, and compared with a traditional microscopic imaging method, the method has higher sensitivity. The invention has the advantages of simple structure, high sensitivity and accurate measurement. The invention can be used for high-resolution three-dimensional microscopic imaging of biological cells, and can be widely applied to nondestructive, unmarked and non-contact three-dimensional tomography of optical transparent objects and the like.
Description
(I) technical field
The invention relates to a novel common-light-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method, which can be used for high-resolution three-dimensional microscopic imaging of biological cells, can be widely applied to nondestructive, unmarked and non-contact three-dimensional tomography of optical transparent objects and the like, and belongs to the technical field of microscopic imaging.
(II) background of the invention
The three-dimensional absorption rate distribution of the cells to be detected is an important inherent property, and for the optically transparent cells to be detected, the three-dimensional absorption rate distribution can reflect the information of the microstructure, the density and the like of a sample, so that nondestructive, label-free and non-contact three-dimensional tomography is required to be realized.
In modern life science research, fluorescence imaging is commonly used to label samples to be tested. However, in the process of labeling, the sample to be tested will be affected to some extent, and the final research result will be affected. The digital holographic tomography technology is a novel imaging technology which is lossless, unmarked and non-contact, can reconstruct and obtain three-dimensional absorption rate distribution information of cells to be detected, and is a research hotspot in recent years.
The digital holographic tomography technology combines the digital holographic microscopic imaging technology and the computed tomography technology, and is a new technology proposed in recent years. In recent years, various imaging methods using digital holography tomography have been proposed, but most of the ideas are to perform digital holography recording by combining with a mach-zehnder interference optical path.
More devices are used for Mach-Zehnder interference optical path imaging, the requirement on system stability is higher, and the operation becomes complicated. The Mach-Zehnder interference light path-based method has higher requirements on devices, and the light path is more complex and difficult to debug, so that a new imaging method is urgently needed, and the method has fewer used devices, simpler light path, higher system stability, simpler and more convenient operation, higher sensitivity for measuring cells to be measured and higher imaging resolution.
The invention provides a novel method for three-dimensional testing of phase-enhanced cell absorption rate of a common-path F-P cavity. In principle, the light beam is reflected for multiple times in the F-P cavity and passes through the cell to be measured for multiple times, the cell to be measured absorbs the light wave for multiple times, the light intensity attenuation is accumulated in sequence, and the absorption is enhanced, so the measurement sensitivity is higher. The hologram formed by the F-P cavity interference has higher fineness than Mach-Zehnder interference, and the recording of the hologram is more accurate.
Patent CN201310082100.9 discloses a digital holographic imaging on-line reconstruction display system and method, which is characterized in that a Mach-Zehnder interference digital holographic recording optical path is adopted, compared with the novel common optical path F-P cavity phase enhanced cell absorption rate three-dimensional test method provided by the invention, the novel method provided by the invention has higher measurement sensitivity.
Patent CN201610911993.7 discloses a dual-wavelength phase microscopic imaging system and method, and a corresponding phase recovery method, which is characterized in that a mach-zehnder interference optical path is adopted to realize a dual-wavelength coaxial phase-shift interference microscopic system.
Patent CN201710518263.5 discloses a three-dimensional refractive index tomography microscopic imaging system and method thereof, which can restore the three-dimensional refractive index information of a sample, but in the imaging optical path, there is an essential difference from the present invention, and the required devices are more complex, and it is also difficult to obtain the three-dimensional absorption rate distribution of the cells to be measured. .
Patent CN201710904860.1 discloses an optical coherence tomography imaging system, which adopts mach-zehnder interference optical path, and is characterized in that optical fiber is adopted to simplify the system and reduce the cost, but the optical path structure is still more complex compared with the optical path structure of F-P cavity.
Patent CN201810145657.5 discloses a high resolution digital holographic diffraction tomography, which is characterized in that a mach-zehnder interference optical path structure is adopted, and a synthetic aperture method is used to obtain N synthetic high resolution holograms, thereby obtaining a high resolution three-dimensional refractive index representation of a measured sample. The structure is relatively more complex, and the invention is essentially different from the invention.
Patent cn201910136421.x discloses a super-resolution digital holographic imaging system and an imaging method, and the imaging system is characterized in that a transmission-type spatial light modulator is added in front of a traditional mach-zehnder interference optical path to modulate a light source. Compared with the invention adopting the optical path structure of the F-P cavity, the invention has essential difference.
The invention discloses a novel three-dimensional test method for phase-enhanced cell absorption rate of a common-light-path F-P cavity, which can be used for high-resolution microscopic imaging of biological cells and can be widely applied to nondestructive, unmarked and non-contact chromatographic imaging of optical transparent objects and the like. The novel method for three-dimensional testing of the phase-enhanced cell absorption rate of the common-light-path F-P cavity adopts a digital holographic recording light path of the common-light-path F-P cavity, and adopts a digital holographic tomography technology to perform chromatographic reconstruction on a digital hologram recorded by the F-P cavity light path so as to restore the three-dimensional absorption rate distribution of cells to be tested. Compared with the prior art, the digital holographic recording light path of the common-light-path F-P cavity is adopted, so that the light beam is reflected in the F-P cavity for multiple times to pass through the cell to be measured, and the measurement sensitivity is higher. The novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method has the advantages of simple structure, high sensitivity and higher resolution.
Disclosure of the invention
The invention aims to provide a novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method which is simple in structure, high in sensitivity and higher in resolution.
The purpose of the invention is realized as follows:
the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method comprises digital hologram recording, numerical reconstruction, error processing and three-dimensional absorption rate distribution reconstruction based on an F-P cavity. Recording a digital hologram when a cell sample to be detected is not put in, then putting a cell to be detected in the F-P cavity, recording an optimal digital hologram containing information of the cell sample to be detected when theta is 0, and carrying out numerical reconstruction on the recorded digital hologram to obtain complex amplitude distribution U of transmitted lightTiThe complex amplitude distribution of the transmitted light is multiplied by the conjugate of the complex amplitude distribution of the transmitted light to obtain the light intensity distribution of the transmitted light, and the light intensity distribution I of the cell sample not to be detected is not put in0Subtracting the light intensity distribution I containing the information of the cell sample to be measuredTiObtaining the absorption light intensity distribution delta I only containing the information of the cell sample to be detected, and obtaining the absorption rate distribution A at the angle according to a derivation formulai. And rotating the cell sample to be detected, recording the optimal digital holograms at different angles, and sequentially obtaining the absorption rate distribution only containing the information of the cell sample to be detected at the angle. The irradon transform is performed on the absorption profile at all angles,the high-resolution three-dimensional absorption rate distribution A of the cells to be detected can be reconstructed.
The novel method for three-dimensional testing of phase-enhanced cell absorption rate of common-path F-P cavity provided by the invention is suitable for a measurement system of a digital holographic recording light path, cells to be tested, a control module and a calculation display module based on the common-path F-P cavity, as shown in figure 2 a. The digital holographic recording light path of the common light path F-P cavity comprises a light source, a beam expander, the F-P cavity, a microscope objective, an image collector and the like, and all devices share a common light path. The cells to be detected are optical fibers with various microstructures. The control module consists of a computer, an instrument control unit and an instrument control interface, and is used for controlling and operating the image collector, the rotary control platform and the like to complete the recording of the digital hologram containing the cell information to be detected. And the calculation display module is used for carrying out program processing on the recorded digital hologram and displaying the three-dimensional absorption rate information of the cell to be detected on line.
The digital holographic recording light path 11 of the common light path F-P cavity comprises a light source, a beam expander, the F-P cavity, a microscope objective, an image collector and the like, and all devices share a common light path. Preferably, the light source is a laser light source with a wavelength of 532nm, a beam expander with a corresponding wavelength is selected, an F-P etalon with a fixed cavity length is selected, a microscope objective with a magnification of 20 times is selected, and devices such as a Charge Coupled Device (CCD) with a pixel size of 3.45 μm × 3.45 μm are selected.
The cell 12 to be measured is located in the F-P cavity of the digital holographic recording light path 11 of the common light path F-P cavity, the light beam is reflected for multiple times in the F-P cavity and passes through the cell to be measured for multiple times, the cell to be measured absorbs the light wave for multiple times, the attenuation of the light intensity is accumulated in sequence, and the absorption is enhanced, so the measurement sensitivity is higher.
In the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method, the cavity length of the F-P cavity is larger than the diameter of the cell 12 to be tested, so that the cell 12 to be tested can be placed in the F-P cavity. The cell 12 to be detected is controlled to rotate by a rotary control platform, and the rotary control platform is an optical tweezers system and the like.
In the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method, the interface reflectivity of an F-P cavity is 0-1.
In the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method, the F-P cavities have cavity lengths with different lengths and can have different shapes and sizes. The cavity length of the F-P cavity may be adjustable, i.e., an F-P etalon, when the cavity length of the F-P cavity is fixed.
The cell 12 to be detected is an optically transparent biological cell. Since the cell 12 to be measured is a minute object and the light beam passes through the cell 12 to be measured, a cylindrical lens effect is generated, and therefore, an absorption rate matching fluid having an absorption rate equivalent to that of the outermost layer of the cell 12 to be measured needs to be selected and filled. The cells to be tested 12 may be biological cells, minute organisms, etc. having various sizes and shapes.
The control module 13 is composed of a computer, an instrument control unit and an instrument control interface, and controls and operates the image collector, the rotary control platform and the like to complete the recording of the digital hologram containing the cell information to be detected. After the digital holographic recording light path 11 of the common light path F-P cavity is built, the control module 13 is utilized to control the rotary control platform to drive the cell 12 to be detected to rotate for a circle, and the CCD is controlled to collect and store the digital hologram containing the information of the cell 12 to be detected.
And the calculation and display module 14 is used for carrying out program processing on the stored digital hologram and displaying the absorption rate information of the cell 12 to be detected on line. According to the novel microscopic imaging method provided by the invention, the digital hologram acquired by the CCD is processed and reconstructed to obtain the high-resolution three-dimensional absorption rate distribution of the cells 12 to be detected.
In the digital holographic recording optical path 11 of the common optical path F-P cavity, the light beam is reflected and transmitted in the F-P cavity for multiple times, as shown in fig. 3b, passes through the cell 12 to be detected for multiple times, the three-dimensional information of the cell 12 to be detected is recorded, the cell to be detected absorbs the light wave for multiple times, the light intensity attenuation is accumulated in sequence, the absorption is enhanced, and the complex amplitude of the light beam finally penetrating through the F-P cavity is:
wherein, UTFor complex amplitude of transmitted light, U is the complex amplitude of incident light in the F-P cavity, and R is the F-P cavityThe surface reflectivities of the inner sides of the two parallel flat glass plates, 31 and 32 in fig. 3a, δ is the phase difference distribution of the cells to be measured.
The intensity distribution of the transmitted light is:
I＝UT·UT *(2)
subtracting the light intensity distribution of the cells to be detected from the light intensity distribution of the cells not to be detected, namely the light intensity distribution absorbed by the cells to be detected, wherein delta I is I0-I. Wherein, I0＝U0·U0 *Light intensity distribution of cells not to be measured, U0The complex amplitude distribution of the cells not put in the test sample.
The ratio of the light intensity distribution absorbed by the cell to be detected to the light intensity distribution without the cell to be detected, namely the absorption rate distribution of the cell to be detected at the angle is as follows:
A＝ΔI/I0(3)
for a common digital holographic recording light path based on Mach-Zehnder interference, a light source is divided into two beams by a beam splitter prism, one beam of light passes through a cell to be detected to serve as object light waves, and the other beam of light serves as reference light. The two beams of light are combined by the beam splitter prism and meet and interfere with each other on the CCD recording plane to form a hologram. The light beam only passes through the cell to be detected once, the three-dimensional information of the cell to be detected is recorded, and the cell to be detected only absorbs the light intensity once. The digital hologram recorded by CCD is reconstructed numerically and the complex amplitude distribution of the reconstructed object light wave is UMZ. Meanwhile, the light intensity distribution of the cells to be detected is obtained as follows:
IMZ＝UMZ·UMZ *(4)
wherein, UMZShowing the complex amplitude distribution of the reproduced object light wave.
The digital holographic recording light path based on Mach-Zehnder interference is characterized in that the light intensity distribution absorbed by the cells to be detected is delta I by subtracting the light intensity distribution of the cells to be detected from the light intensity distribution of the cells not to be detectedMZ＝IMZ0-IMZ。
The digital holographic recording light path based on Mach-Zehnder interference has light beam passing through cell to be measured along propagation directionAfter passing through the cell to be detected once, the light intensity absorbed by the cell to be detected is accumulated once along the light beam transmission direction. As can be seen from FIG. 3b, in the common-path F-P cavity, the incident beam is 33, the incident angle is 34, and the beam is reflected and transmitted in the F-P cavity for multiple times and passes through the cell to be detected for multiple times. When the incident beam 33 vertically enters, the light beam is reflected and transmitted for multiple times in the F-P cavity, passes through the cell to be detected for multiple times, the three-dimensional information of the cell to be detected is recorded, the cell to be detected absorbs the light wave for multiple times, the attenuation of the light intensity is accumulated in sequence, and the absorption is enhanced, namely delta IMZ< Δ I. Therefore, the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method is higher in measurement sensitivity, higher in resolution and superior to a common absorption rate test method based on Mach-Zehnder interference in principle.
Therefore, in the digital hologram recording optical path 11 of the common optical path F-P cavity, a digital hologram when the sample of the cell 12 to be measured is not put is recorded, then the cell 12 to be measured is put in the F-P cavity, when θ is 0, an optimal digital hologram including the sample information of the cell 121 to be measured is recorded, and the recorded digital hologram is numerically reconstructed to obtain the complex amplitude distribution U of the transmitted lightTiThe complex amplitude distribution of the transmitted light is multiplied by the conjugate of the complex amplitude distribution of the transmitted light, so that the light intensity distribution of the transmitted light can be obtained, and the light intensity distribution I of the sample without the cell 12 to be detected is obtained0Light intensity distribution I obtained by subtracting information on sample containing cells 12 to be measuredTiObtaining the absorption light intensity distribution delta I only containing the information of the cell 12 sample to be detected, from Ai＝ΔI/I0The absorption profile A at this angle can be obtainedi. And rotating the cell 12 sample to be detected, recording the optimal digital holograms at different angles, and sequentially obtaining the absorption rate distribution only containing the information of the cell 12 sample to be detected at the angle. And performing iRadon transformation on the absorption rate distribution at all angles to reconstruct the high-resolution three-dimensional absorption rate distribution A of the cell 12 to be detected.
The invention relates to a novel three-dimensional test method for phase-enhanced cell absorption rate of a common-path F-P cavity, which mainly comprises the following steps:
the first step is as follows: and recording the digital hologram when the F-P cavity is not filled with the cells to be detected. The F-P cavity is filled with the cells to be tested.
The second step is that: the control module controls the rotary control platform to rotate the cell to be detected, records the optimal digital hologram after the cell to be detected is placed at the angle, and intercepts a certain size.
The third step: the recorded digital hologram is numerically reconstructed to obtain a complex amplitude distribution of the transmitted light at the angle, which is UTi。
The fourth step: the light intensity distribution I obtained by not putting the cells to be detected0Subtracting the light intensity distribution I obtained by placing the cell to be measuredTiAnd obtaining the light intensity distribution delta I absorbed by the cells to be detected, wherein the angle only contains the information of the cells to be detected.
The fifth step: according to formula Ai＝ΔI/I0The absorption rate distribution of the cells to be detected at the angle can be obtained.
And a sixth step: and the control module controls the rotary control platform to rotate the cell to be detected for a circle, and the second step to the fifth step are repeated in sequence to obtain the absorption rate distribution of the cell to be detected at different angles.
The seventh step: and through the calculation and display module, the absorption rate distribution of the cells to be detected with all angles on different sections is subjected to iRadon transformation, and the high-resolution three-dimensional absorption rate distribution A (x, y, z) of the cells to be detected can be displayed on line.
The invention provides a novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method which comprises digital hologram recording, numerical reconstruction, error processing and three-dimensional absorption rate distribution reconstruction based on an F-P cavity. Compared with the prior art, the digital holographic recording light path of the common-light-path F-P cavity is adopted, so that the light beam is reflected in the F-P cavity for multiple times to pass through the cell to be measured, and the measurement sensitivity is higher. The novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method has the advantages of simple structure, high sensitivity and higher resolution.
(IV) description of the drawings
Fig. 1 is a schematic structural diagram of a novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method. Including digital hologram recording, numerical reconstruction, error processing, and three-dimensional absorptivity distribution reconstruction based on the F-P cavity.
Fig. 2a is a structural diagram of a measurement system applicable to the novel common-path F-P cavity phase-enhanced cell absorption rate three-dimensional test method provided by the invention. The device comprises a digital holographic recording light path 11 of a common light path F-P cavity, a cell 12 to be detected, a control module 13 and a calculation display module 14. FIG. 2b is a schematic diagram of a digital holographic recording optical path of a common optical path F-P cavity in an embodiment of the present invention. In this embodiment, a digital holographic recording optical path of a common optical path F-P cavity includes a light source 21, an attenuator 22, a beam expander 23, an F-P etalon 24, a cell to be measured 25, a microscope objective lens 26, a CCD27, and a computer 28.
Figure 3a is a schematic view of an F-P chamber of the present invention. The F-P cavity consists of two parallel planar glass plates with dielectric films of reflectivity R plated on their inward facing sides 31 and 32. Fig. 3b is a schematic diagram of the light beam incident on the F-P cavity, reflected multiple times in the F-P cavity, and transmitted. 33 is an incident light wave, 34 is an incident angle, 35, 37, 39 are schematic diagrams of transmitted light, and 36, 38 are schematic diagrams of reflected light.
FIG. 4 is a schematic diagram of the F-P chamber and the test cells in an embodiment of the invention. The rotating control platform 41 controls the cell 25 to be detected to rotate, the cell 25 to be detected is placed in the cuvette 42, the cuvette 42 is filled with refractive index matching liquid with the refractive index equivalent to that of the outermost layer of the cell 25 to be detected, and the propagation direction of the light beam is shown as 43.
FIG. 5 is a flow chart of the steps of the novel method for three-dimensional measurement of phase-enhanced cell absorption rate of common-path F-P cavity according to the present invention.
(V) detailed description of the preferred embodiments
The invention is further illustrated below with reference to specific examples.
Example 1:
FIG. 2b shows an embodiment of a common optical path F-P cavity digital holographic recording optical path. The light path comprises a light source 21, an attenuator 22, a beam expander 23, an F-P etalon 24, a cell to be detected 25, a microscope objective lens 26, a CCD27 and a computer 28. The light beam starts from a light source 21, passes through an attenuator 22, the energy of the light beam is weakened, the light beam passes through a beam expander 23, the diameter of the light beam is enlarged, when the light beam passes through an F-P etalon 24 containing a cell 25 to be detected, the light beam is reflected for multiple times in the F-P etalon 24, passes through the cell 25 to be detected for multiple times, each transmitted light beam passing through the F-P etalon 24 passes through a microscope objective lens 26, is subjected to interference superposition on a CCD27, and is recorded with an interference digital hologram by a CCD27 and is stored in a computer 28.
In the present embodiment, preferably, the light source 21 is a laser light source with a wavelength of 532nm, and selects a corresponding wavelength beam expander 23, selects an adjustable attenuator 22, selects an F-P etalon 24 with a cavity length of 60mm, selects a microscope objective lens 26 with a magnification of 20 times, and selects devices such as a CCD27 with a pixel size of 3.45 μm × 3.45 μm.
In this embodiment, preferably, as shown in fig. 3a, the F-P etalon 24 is formed by two parallel flat glass plates coated with a dielectric film with a reflectivity of 0.9 on the inner surface to form the F-P etalon 24 with a cavity length of 60 mm. Preferably, the cell 25 to be detected is paramecium, the paramecium is placed in the cuvette 42, and the cuvette 42 is filled with refractive index matching fluid with the refractive index equivalent to that of the outermost layer of the paramecium. The cuvette 42 is placed in the cavity of the F-P etalon 24 and the digital holographic recording light path is adjusted so that the optical axis of the light beam 43 passes through the cell 25 to be measured as shown in fig. 4.
In this embodiment, preferably, the rotating control platform 41 is an optical tweezers system, and the control module precisely controls the cell 25 to be measured to rotate around the axial direction of the rotating shaft by one rotation.
In this embodiment, the digital holographic recording optical path of the common optical path F-P cavity includes a light source 21, an attenuator 22, a beam expander 23, an F-P etalon 24, a microscope objective lens 26, a CCD27, and the like. The cell 25 to be detected is paramecium. The control module 3 is composed of a computer 28, an instrument control unit and an instrument control interface, and controls and operates the CCD27, the rotary control platform 41 for controlling the rotation of the cell 25 to be detected, and the like, to complete the recording of the digital hologram including the information of the cell 25 to be detected. And the calculation display module 4 is used for carrying out program processing on the recorded digital hologram according to the microscopic imaging method provided by the invention and displaying the three-dimensional absorption rate information of the cell to be detected on line.
In this embodiment, a new method for three-dimensional measurement of phase-enhanced cell absorption rate in a common optical path F-P cavity is used to process a digital hologram of a cell 25 to be measured, and to display three-dimensional absorption rate information of the cell 25 to be measured on line, as shown in fig. 5, the method mainly includes the following steps:
first step 51: a digital hologram is recorded of the F-P etalon 24 without the test cell 25 placed therein. The test cell 25 is placed in the F-P etalon 24.
Second step 52: the control module controls the optical tweezers system to enable the cell 25 to be detected to rotate around the shaft, records the optimal digital hologram after the cell 25 to be detected is placed at the angle, and intercepts the optimal digital hologram with a certain size.
Third step 53: the recorded digital hologram is reconstructed numerically.
The fourth step 54: and obtaining the light intensity distribution containing the information of the sample of the cell 25 to be detected at the angle according to the formula (2).
Fifth step 55: the light intensity distribution I obtained without placing the cell 25 to be measured0Subtracting the light intensity distribution I obtained by placing the cell 25 to be measuredTiAnd obtaining the light intensity distribution delta I absorbed by the cell 25 to be detected, wherein the angle only contains the sample information of the cell 25 to be detected.
Sixth step 56: according to the formula (3), the absorption rate distribution of the test cell 25 at the angle can be obtained.
Seventh step 57: the control module controls the optical tweezers system to rotate once every 1 degree, so that the cell 25 to be detected rotates for a circle, and the second step 52 to the sixth step 56 are sequentially repeated, so that the absorption rate distribution of the cell 25 to be detected at different angles can be obtained.
Eighth step 58: by calculating the display module 4, the absorbance distribution of the cell 25 to be detected on the cross section at all angles is subjected to iRadon transformation, and the reconstructed high-resolution three-dimensional absorbance distribution a (x, y, z) of the cell 25 to be detected can be displayed on line.
The above description is only a preferred embodiment of the present invention, but the scope of the present invention is not limited thereto. Various modifications and alterations of this invention will occur to those skilled in the art in view of the spirit and scope of this invention and are intended to be encompassed by the following claims.
Claims (7)
1. A novel three-dimensional test method for the common-path F-P cavity enhanced cell absorption rate. The method is characterized in that:including digital hologram recording, numerical reconstruction, error processing, and three-dimensional absorptivity distribution reconstruction based on the F-P cavity. Recording a digital hologram when a cell sample to be detected is not put in, then putting a cell to be detected in the F-P cavity, recording an optimal digital hologram containing information of the cell sample to be detected when theta is 0, and carrying out numerical reconstruction on the recorded digital hologram to obtain complex amplitude distribution U of transmitted lightTiThe complex amplitude distribution of the transmitted light is multiplied by the conjugate of the complex amplitude distribution of the transmitted light to obtain the light intensity distribution of the transmitted light, and the light intensity distribution I of the cell sample not to be detected is not put in0Subtracting the light intensity distribution I containing the information of the cell sample to be measuredTiObtaining the absorption light intensity distribution delta I only containing the information of the cell sample to be detected, and obtaining the absorption rate distribution A at the angle according to a derivation formulai. And rotating the cell sample to be detected, recording the optimal digital holograms at different angles, and sequentially obtaining the absorption rate distribution only containing the information of the cell sample to be detected at the angle. And performing iRadon transformation on the absorption rate distribution at all angles to reconstruct the high-resolution three-dimensional absorption rate distribution A of the cells to be detected.
2. A novel three-dimensional test method for the common-path F-P cavity enhanced cell absorption rate. The method is characterized in that: the measuring system is suitable for a digital holographic recording light path based on a common light path F-P cavity, cells to be measured, a control module and a calculation display module. The digital holographic recording light path of the common light path F-P cavity comprises a light source, a beam expander, the F-P cavity, a microscope objective, an image collector and the like, and all devices share a common light path. The cells to be detected are optical fibers with various microstructures. The control module consists of a computer, an instrument control unit and an instrument control interface, and is used for controlling and operating the image collector, the rotary control platform and the like to complete the recording of the digital hologram containing the cell information to be detected. And the calculation display module is used for carrying out program processing on the recorded digital hologram and displaying the three-dimensional absorption rate information of the cell to be detected on line.
3. The novel method for three-dimensional measurement of the absorption rate of the common-path F-P cavity enhanced cell as claimed in claim 2, wherein the cell to be measured is located in the F-P cavity in the digital holographic recording optical path 1.
4. The novel common-path F-P cavity enhanced cell absorption rate three-dimensional test method according to claim 2, wherein the cavity length of the F-P cavity is larger than the diameter of a cell to be tested, and the interface reflectivity of the F-P cavity is 0-1.
5. The novel three-dimensional measurement method for the enhanced cellular absorptivity of the common-path F-P cavity according to claim 2, wherein the F-P cavity has cavity lengths with different lengths.
6. The novel method for three-dimensional measurement of the absorption rate of the common-path F-P cavity-enhanced cells according to claim 1, wherein the cells to be measured can be biological cells, micro organisms and the like with different sizes and shapes.
7. The novel method for three-dimensional measurement of the enhanced cellular absorptivity of the common-path F-P cavity according to claim 1, which mainly comprises the following steps:
the first step is as follows: and recording the digital hologram when the F-P cavity is not filled with the cells to be detected. The F-P cavity is filled with the cells to be tested.
The second step is that: the control module controls the rotary control platform to rotate the cell to be detected, records the optimal digital hologram after the cell to be detected is placed at the angle, and intercepts a certain size.
The third step: the recorded digital hologram is numerically reconstructed to obtain a complex amplitude distribution of the transmitted light at the angle, which is UTi。
The fourth step: the light intensity distribution I obtained by not putting the cells to be detected0Subtracting the light intensity distribution I obtained by placing the cell to be measuredTiAnd obtaining the light intensity distribution delta I absorbed by the cells to be detected, wherein the angle only contains the information of the cells to be detected.
The fifth step: according to formula Ai＝ΔI/I0The absorption rate distribution of the cells to be detected at the angle can be obtained.
And a sixth step: and the control module controls the rotary control platform to rotate the cell to be detected for a circle, and the second step to the fifth step are repeated in sequence to obtain the absorption rate distribution of the cell to be detected at different angles.
The seventh step: and through the calculation and display module, the absorption rate distribution of the cells to be detected with all angles on different sections is subjected to iRadon transformation, and the high-resolution three-dimensional absorption rate distribution A (x, y, z) of the cells to be detected can be displayed on line.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911089768.XA CN111122446A (en) | 2019-11-08 | 2019-11-08 | Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911089768.XA CN111122446A (en) | 2019-11-08 | 2019-11-08 | Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111122446A true CN111122446A (en) | 2020-05-08 |
Family
ID=70495771
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201911089768.XA Pending CN111122446A (en) | 2019-11-08 | 2019-11-08 | Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111122446A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111796501A (en) * | 2020-06-28 | 2020-10-20 | 西安工业大学 | Common-path phase shift digital holographic microscopic measuring device |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080239428A1 (en) * | 2007-04-02 | 2008-10-02 | Inphase Technologies, Inc. | Non-ft plane angular filters |
| EP2327955A1 (en) * | 2008-09-05 | 2011-06-01 | Universidad Politécnica De Madrid | Optical detection system for labelling-free high-sensitivity bioassays |
| US20120116703A1 (en) * | 2009-04-24 | 2012-05-10 | Nicolas Pavillon | Method and apparatus for enhanced spatial bandwidth wavefronts reconstructed from digital interferograms or holograms |
| US20180266806A1 (en) * | 2017-03-17 | 2018-09-20 | National Taiwan Normal University | Method and apparatus for digital holographic microtomography |
| CN110366678A (en) * | 2016-12-27 | 2019-10-22 | 密执安州立大学董事会 | Microscope based on Laser emission |
-
2019
- 2019-11-08 CN CN201911089768.XA patent/CN111122446A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080239428A1 (en) * | 2007-04-02 | 2008-10-02 | Inphase Technologies, Inc. | Non-ft plane angular filters |
| EP2327955A1 (en) * | 2008-09-05 | 2011-06-01 | Universidad Politécnica De Madrid | Optical detection system for labelling-free high-sensitivity bioassays |
| US20120116703A1 (en) * | 2009-04-24 | 2012-05-10 | Nicolas Pavillon | Method and apparatus for enhanced spatial bandwidth wavefronts reconstructed from digital interferograms or holograms |
| CN110366678A (en) * | 2016-12-27 | 2019-10-22 | 密执安州立大学董事会 | Microscope based on Laser emission |
| US20180266806A1 (en) * | 2017-03-17 | 2018-09-20 | National Taiwan Normal University | Method and apparatus for digital holographic microtomography |
Non-Patent Citations (2)
| Title |
|---|
| 欧阳丽婷等: "老鼠大脑海马区神经元活细胞的数字全息相衬成像实验研究", 《中国激光》 * |
| 王华英等: "高分辨率数字全息相衬成像技术", 《深圳大学学报(理工版)》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111796501A (en) * | 2020-06-28 | 2020-10-20 | 西安工业大学 | Common-path phase shift digital holographic microscopic measuring device |
| CN111796501B (en) * | 2020-06-28 | 2022-03-22 | 西安工业大学 | Common-path phase shift digital holographic microscopic measuring device |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102540446B (en) | High-speed structure illumination optical microscope system and method based on digital micromirror device | |
| US11781966B2 (en) | 3D diffraction tomography microscopy imaging method based on LED array coded illumination | |
| US9546952B2 (en) | Distribution of refractive index measurement by synthetic aperture tomography | |
| Haeberlé et al. | Tomographic diffractive microscopy: basics, techniques and perspectives | |
| US7880891B1 (en) | Total internal reflection holographic microscope | |
| CN110455799B (en) | High-resolution holographic microscope and method for live cell imaging | |
| CN108971747A (en) | A kind of ultrafast laser micro-nano technology device having on-line monitoring function | |
| JP2005512127A (en) | Apparatus and method for digital holographic imaging | |
| CN104062233A (en) | Precise surface defect scattering three-dimensional microscopy imaging device | |
| Yan et al. | 3D refractive index measurements of special optical fibers | |
| CN106123770B (en) | A kind of method that refractive index dynamically measures simultaneously with pattern | |
| CN108918465A (en) | Optical 3-dimensional imaging system and optical 3-dimensional imaging method | |
| CN208580026U (en) | Optical 3-dimensional imaging system | |
| CN111122446A (en) | Novel three-dimensional test method for phase-enhanced cell absorption rate of common-path F-P cavity | |
| CN111289479B (en) | Phase defect detection device and method based on nonlinear thermography inversion | |
| CN111025874B (en) | Novel phase-shift phase microscopic imaging method based on F-P cavity | |
| CN110987871A (en) | Novel multi-wavelength phase microscopic imaging method based on F-P cavity | |
| CN115523865A (en) | Three-dimensional measuring device and measuring method for laser damage of optical element | |
| AU2020103535A4 (en) | A new cell absorption rate three-dimensional (3D) testing method enhanced by a common path F-P cavity | |
| US20230324275A1 (en) | A simple in-line digital holography system for measuring 3d cell shape | |
| AU2020103534A4 (en) | A testing method for high-resolution three-dimensional (3D) refractive index of microstructured optical fiber | |
| CN111198169A (en) | Microstructure optical fiber high resolution three-dimensional refractive index testing method | |
| CN111982014B (en) | Micro-interference-based microsphere surface morphology large-field-of-view measurement method | |
| AU2020103536A4 (en) | A new phase-shifting phase micro imaging method based on an F-P cavity | |
| AU2020103537A4 (en) | A new multiwavelength phase micro imaging method based on an F-P cavity |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20200508 |
|
| WD01 | Invention patent application deemed withdrawn after publication |