CN113916776A - Axial positioning system and method based on virtual pinhole confocal technology - Google Patents
Axial positioning system and method based on virtual pinhole confocal technology Download PDFInfo
- Publication number
- CN113916776A CN113916776A CN202110992111.5A CN202110992111A CN113916776A CN 113916776 A CN113916776 A CN 113916776A CN 202110992111 A CN202110992111 A CN 202110992111A CN 113916776 A CN113916776 A CN 113916776A
- Authority
- CN
- China
- Prior art keywords
- spectroscope
- sample
- axial positioning
- light source
- lens
- 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
- 238000005516 engineering process Methods 0.000 title claims abstract description 17
- 238000000034 method Methods 0.000 title claims description 26
- 230000003287 optical effect Effects 0.000 claims abstract description 16
- 238000001514 detection method Methods 0.000 abstract description 12
- 101100327165 Arabidopsis thaliana CCD8 gene Proteins 0.000 description 7
- 239000000835 fiber Substances 0.000 description 4
- 238000005286 illumination Methods 0.000 description 4
- 238000003384 imaging method Methods 0.000 description 3
- 238000010586 diagram Methods 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000004624 confocal microscopy Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 238000000386 microscopy Methods 0.000 description 1
- 238000003325 tomography 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
- G01N21/01—Arrangements or apparatus for facilitating the optical investigation
-
- 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
- G01N21/84—Systems specially adapted for particular applications
-
- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/0004—Microscopes specially adapted for specific applications
- G02B21/002—Scanning microscopes
- G02B21/0024—Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
- G02B21/0032—Optical details of illumination, e.g. light-sources, pinholes, beam splitters, slits, fibers
-
- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/0004—Microscopes specially adapted for specific applications
- G02B21/002—Scanning microscopes
- G02B21/0024—Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
- G02B21/008—Details of detection or image processing, including general computer control
Abstract
The invention discloses an axial positioning system based on a virtual pinhole confocal technology, which comprises: the confocal detection device comprises an incoherent light source, a collimating lens, a spectroscope, an objective lens, a sample stage, a focusing lens and a photoelectric detector CCD, wherein the sample stage, the objective lens, the spectroscope, the focusing lens and the photoelectric detector CCD are positioned on the same longitudinal optical axis, the incoherent light source and the collimating lens are positioned on the same horizontal optical axis, the incoherent light source and the collimating lens are sequentially arranged on an incident light path of the spectroscope, the objective lens and the sample stage are sequentially arranged on one side of the spectroscope, which is far away from the objective lens, and the focusing lens and the photoelectric detector CCD are sequentially arranged on a reflected light path of the spectroscope.
Description
Technical Field
The invention relates to an axial positioning system and method based on a virtual pinhole confocal technology, and belongs to the technical field of optical measurement.
Background
The confocal microscope is invented by M.Minsky after doctor of Harvard university in America at the middle and later stages of the 50 th century, and is a device for realizing three-dimensional imaging of an internal three-dimensional structure of a sample by adopting point illumination and point detection and introducing point-by-point scanning. The confocal microscopy device has high transverse resolution characteristics and unique optical tomography capability, and has wide influence on a plurality of fields. The invention and application of the confocal microscopic measuring device start the research and application of the optical chromatography microscopy. The method breaks through the limit of optical diffraction resolution, is a hotspot and difficult problem of international research for a long time, and has emerged various solutions in the time of nearly twenty years from the end of the 20 th century to the beginning of the 21 st century, common confocal detectors comprise pinholes, slits, single-mode fibers, multi-mode fibers, annular holes, square holes, virtual pinholes and the like, fixed pinhole filtering confocal detection is adopted, such as physical pinholes, single-mode fibers and multi-mode fibers, the imaging resolution and the optical chromatographic capacity of a system are determined after the geometric shape is determined, and the size of the detection pinhole is selected by methods such as electric control of a precise pinhole turntable and the like in practical application, so that the compromise of the resolution, the chromatographic characteristic and the signal-to-noise ratio of the system is realized; transverse super-resolution imaging can be realized by adopting large and small pinhole filtering differential confocal detection.
The fixed-size pinhole limits flexible adjustment of resolution on the one hand and needs to be realized by double-path confocal detection on the light path on the other hand. In addition, confocal microscope systems achieve confocal detection using a combination of point detection elements and photodetectors, and there is no separate, integrated confocal detector. The above method has a complicated structure and is difficult to implement, so that a simple method is required for implementing confocal detection.
Disclosure of Invention
The invention aims to overcome the defects in the prior art, provides an axial positioning method based on a virtual pinhole confocal technology, has a simple structure and low cost, and obviously reduces the difficulty of realizing confocal detection.
In order to achieve the purpose, the invention is realized by adopting the following technical scheme:
in a first aspect, the present invention provides an axial positioning system based on a virtual pinhole confocal technique, including: incoherent light source, collimating lens, spectroscope, objective, sample platform, focusing lens and photoelectric detector CCD, sample platform, objective, spectroscope, focusing lens, photoelectric detector CCD are located same longitudinal optical axis, incoherent light source, collimating lens are located same horizontal optical axis, wherein, incoherent light source, collimating lens set up in order on the incident light path of spectroscope, objective, sample platform set up in order on the reflection light path of spectroscope, focusing lens, photoelectric detector CCD set up in order on the spectroscope deviates from one side of objective.
Furthermore, a scanning control platform is arranged at the bottom of the sample platform, and a driving device is arranged on the scanning control platform and used for driving the sample platform to realize stepping on the longitudinal optical axis.
Furthermore, the driving device is driven by an electric push rod or a screw rod.
Furthermore, the scanning control platform is provided with a control system for accurately controlling the driving distance of the driving device.
Furthermore, a control button is arranged on the scanning control platform.
Further, the incoherent light source is an LED light source or a common light source.
In a second aspect, the present invention provides an axial positioning method based on a virtual pinhole confocal technique, where the method is based on any one of the above axial positioning systems, and the method includes:
placing a sample on a sample stage, collimating light emitted by an incoherent light source through a collimating lens, and illuminating the sample through a spectroscope and an objective lens;
the reflected light of the sample is projected to a photoelectric detector CCD photosensitive surface through an objective lens, a spectroscope and a focusing lens;
and taking partial pixels in the middle of the image field of the CCD photosensitive surface of the photoelectric detector as an observation object, and calculating the average intensity value of the partial pixels.
Further, the method further comprises:
driving the sample to step in the axial direction, and calculating the strength value of each step;
and drawing an intensity curve, wherein the peak value of the curve is the focal plane of the adopted objective lens, and the focal plane is used as a reference plane.
Compared with the prior art, the invention has the following beneficial effects:
the invention provides an axial positioning method based on a virtual pinhole confocal technology, which comprises the steps of placing a sample on a sample table, collimating light emitted by an incoherent light source through a collimating lens, and then illuminating the sample through a spectroscope and an objective lens; the reflected light of the sample is projected to a photoelectric detector CCD photosensitive surface through an objective lens, a spectroscope and a focusing lens; and partial pixels in the middle of the image field of the CCD photosensitive surface of the photoelectric detector are taken as observation objects, the average intensity value is calculated, actual pinhole illumination and pinhole detection are not adopted, the structure is effectively simplified, and the cost is obviously reduced.
Drawings
Fig. 1 is a system schematic diagram of an axial positioning system based on a virtual pinhole confocal technology according to an embodiment of the present invention.
In the figure: 1. an incoherent light source; 2. a collimating lens; 3. a beam splitter; 4. an objective lens; 5. a sample stage; 6. scanning the console; 7. a focusing lens; 8. and a photoelectric detector CCD.
Detailed Description
The invention is further described below with reference to the accompanying drawings. The following examples are only for illustrating the technical solutions of the present invention more clearly, and the protection scope of the present invention is not limited thereby.
Example 1
This embodiment introduces an axial positioning system based on virtual pinhole confocal technology, including: incoherent light source 1, collimating lens 2, spectroscope 3, objective 4, sample platform 5, focusing lens 7 and photoelectric detector CCD8, sample platform 5, objective 4, spectroscope 3, focusing lens 7, photoelectric detector CCD8 are located same longitudinal optical axis, incoherent light source 1, collimating lens 2 are located same horizontal optical axis, wherein, incoherent light source 1, collimating lens 2 set up in order on the incident light path of spectroscope 3, objective 4, sample platform 5 set up in order on the reflection light path of spectroscope 3, focusing lens 7, photoelectric detector CCD8 set up in order on the one side that spectroscope 3 deviates from the objective.
The bottom of the sample table 5 is provided with a scanning control table 6, the scanning control table 6 is provided with a driving device for driving the sample table 5 to realize stepping on a longitudinal optical axis, the driving device is driven by an electric push rod or a lead screw, the scanning control table 6 is provided with a control system for accurately controlling the driving distance of the driving device, and the scanning control table 6 is provided with a control button; the incoherent light source 1 is an LED light source or a common light source.
Example 2
The embodiment provides an axial positioning method based on a virtual pinhole confocal technology, where the method is based on the positioning system described in embodiment 1, and the method includes:
a sample is placed on a sample stage 5, light emitted by an incoherent light source 1 is collimated by a collimating lens 2, and then passes through a spectroscope 3 and an objective lens 4 to illuminate the sample;
the reflected light of the sample is projected to a photosensitive surface of a photoelectric detector CCD8 through an objective lens 4, a spectroscope 3 and a focusing lens 7;
the part of pixels in the middle of the photosensitive surface image field of the photoelectric detector CCD8 are taken as an observation object, and the average value of the intensity of the pixels is calculated.
Driving the sample to step in the axial direction, and calculating the strength value of each step;
an intensity curve is drawn, the peak of the curve is the focal plane of the objective 4 used, which is the reference plane.
The application process of the axial positioning method based on the virtual pinhole confocal technology provided by the embodiment specifically relates to the following steps:
fig. 1 shows a schematic diagram of a designed virtual pinhole confocal positioning system, the confocal positioning system adopts an incoherent light source 1 for illumination, a sample is illuminated by an objective 4 after collimation, reflected light of the sample is imaged on a CCD photoelectric detector after passing through the objective 4 and a tube lens, a scanning console 6 drives the sample to realize stepping in the axial direction, simultaneously, one image is acquired every time the sample is axially stepped, and finally, a confocal axial intensity curve is obtained by extracting an average value of intensities of pixel points (such as 10 × 10 pixels) in the same area of the image, thereby realizing accurate positioning of the sample.
The invention provides an axial positioning method based on a virtual pinhole confocal technology, which comprises the steps that a sample is placed on a sample table 5, light emitted by an incoherent light source 1 is collimated by a collimating lens 2, and then passes through a spectroscope 3 and an objective lens 4 to illuminate the sample; the reflected light of the sample is projected to a photosensitive surface of a photoelectric detector CCD8 through an objective lens 4, a spectroscope 3 and a focusing lens 7; and partial pixels in the middle of the photosensitive surface image field of the photoelectric detector CCD8 are taken as observation objects, the average intensity value is calculated, actual pinhole illumination and pinhole detection are not adopted, the structure is effectively simplified, and the cost is obviously reduced.
The above description is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, several modifications and variations can be made without departing from the technical principle of the present invention, and these modifications and variations should also be regarded as the protection scope of the present invention.
Claims (8)
1. An axial positioning system based on virtual pinhole confocal technology, comprising: an incoherent light source (1), a collimating lens (2), a spectroscope (3), an objective lens (4), a sample stage (5), a focusing lens (7) and a photoelectric detector CCD (8),
sample platform (5), objective (4), spectroscope (3), focusing lens (7), photoelectric detector CCD (8) are located same longitudinal optical axis, incoherent light source (1), collimating lens (2) are located same horizontal optical axis, wherein, incoherent light source (1), collimating lens (2) set up in order on the incident light path of spectroscope (3), objective (4), sample platform (5) set up in order on the reflection light path of spectroscope (3), focusing lens (7), photoelectric detector CCD (8) set up in order and deviate from in spectroscope (3) one side of objective (4).
2. The virtual pinhole confocal technology-based axial positioning system of claim 1, wherein: the bottom of sample platform (5) is equipped with scanning control platform (6), be equipped with drive arrangement on scanning control platform (6) for it is step-by-step to drive sample platform (5) and realize on vertical optical axis.
3. The virtual pinhole confocal technique-based axial positioning system of claim 2, wherein: the driving device is driven by an electric push rod or a screw rod.
4. The virtual pinhole confocal technology-based axial positioning system of claim 3, wherein: and the scanning control platform (6) is provided with a control system for accurately controlling the driving distance of the driving device.
5. The virtual pinhole confocal technology-based axial positioning system of claim 4, wherein: and a control button is arranged on the scanning control platform (6).
6. The virtual pinhole confocal technology-based axial positioning system of claim 1, wherein: the incoherent light source (1) is an LED light source or a common light source.
7. An axial positioning method based on a virtual pinhole confocal technology is characterized in that: the method is based on the axial positioning system of any one of claims 1 to 6, and comprises the following steps:
a sample is placed on a sample stage (5), light emitted by an incoherent light source (1) is collimated by a collimating lens (2), and then passes through a spectroscope (3) and an objective lens (4) to illuminate the sample;
reflected light of the sample is projected to a photosensitive surface of a photoelectric detector CCD (8) through an objective lens (4), a spectroscope (3) and a focusing lens (7);
and taking partial pixels in the middle of the photosensitive surface image field of the photoelectric detector CCD (8) as an observation object, and calculating the average intensity value of the partial pixels.
8. The virtual pinhole confocal technique-based axial positioning method of claim 7, further comprising:
driving the sample to step in the axial direction, and calculating the strength value of each step;
and drawing an intensity curve, wherein the peak value of the curve is the focal plane of the adopted objective lens (4), and the focal plane is used as a reference plane.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110992111.5A CN113916776A (en) | 2021-08-27 | 2021-08-27 | Axial positioning system and method based on virtual pinhole confocal technology |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110992111.5A CN113916776A (en) | 2021-08-27 | 2021-08-27 | Axial positioning system and method based on virtual pinhole confocal technology |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113916776A true CN113916776A (en) | 2022-01-11 |
Family
ID=79233223
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110992111.5A Pending CN113916776A (en) | 2021-08-27 | 2021-08-27 | Axial positioning system and method based on virtual pinhole confocal technology |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113916776A (en) |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6548796B1 (en) * | 1999-06-23 | 2003-04-15 | Regents Of The University Of Minnesota | Confocal macroscope |
| CN1971333A (en) * | 2006-10-11 | 2007-05-30 | 南开大学 | Confocal micro imaging system using dummy pinhole |
| CN102768015A (en) * | 2012-07-05 | 2012-11-07 | 哈尔滨工业大学 | Fluorescence response follow-up pinhole microscopic confocal measuring device |
| WO2017000364A1 (en) * | 2015-07-02 | 2017-01-05 | 哈尔滨工业大学 | Probe sensing method and apparatus based on optical beam scanning confocal detection technique |
| CN107121422A (en) * | 2017-06-21 | 2017-09-01 | 中国科学院苏州生物医学工程技术研究所 | A kind of parallel confocal microscopic imaging device and method based on digital micromirror array |
| CN112730266A (en) * | 2020-12-16 | 2021-04-30 | 华中科技大学 | Polarization reflection measurement system and structural parameter measurement method |
-
2021
- 2021-08-27 CN CN202110992111.5A patent/CN113916776A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6548796B1 (en) * | 1999-06-23 | 2003-04-15 | Regents Of The University Of Minnesota | Confocal macroscope |
| CN1971333A (en) * | 2006-10-11 | 2007-05-30 | 南开大学 | Confocal micro imaging system using dummy pinhole |
| CN102768015A (en) * | 2012-07-05 | 2012-11-07 | 哈尔滨工业大学 | Fluorescence response follow-up pinhole microscopic confocal measuring device |
| WO2017000364A1 (en) * | 2015-07-02 | 2017-01-05 | 哈尔滨工业大学 | Probe sensing method and apparatus based on optical beam scanning confocal detection technique |
| CN107121422A (en) * | 2017-06-21 | 2017-09-01 | 中国科学院苏州生物医学工程技术研究所 | A kind of parallel confocal microscopic imaging device and method based on digital micromirror array |
| CN112730266A (en) * | 2020-12-16 | 2021-04-30 | 华中科技大学 | Polarization reflection measurement system and structural parameter measurement method |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US10690898B2 (en) | Light-field microscope with selective-plane illumination | |
| US8081309B2 (en) | Optical microscope and spectrum measuring method | |
| US5754291A (en) | Micro-imaging system | |
| US6084991A (en) | CCD imager for confocal scanning microscopy | |
| CN101303269B (en) | Optical system evaluation apparatus, optical system evaluation method and program thereof | |
| US8472113B2 (en) | Scanning microscope and method for light-microscopic imaging of an object | |
| CN100417931C (en) | Microarray chip detection system | |
| EP0834758A2 (en) | Autofocus system for scanning microscopy | |
| CN107526156A (en) | Mating plate microscope and for running the microscopical method of mating plate | |
| JP2012237647A (en) | Multifocal confocal raman spectroscopic microscope | |
| KR102419163B1 (en) | Real-time autofocus focusing algorithm | |
| JP7009619B2 (en) | Double-pass macro image | |
| JP2010054391A (en) | Optical microscope, and method of displaying color image | |
| CN113589506B (en) | Biological microscopic vision pre-focusing device and method based on spectrum confocal principle | |
| JP2021192118A (en) | Two-dimensional and three-dimensional fixed z scanning | |
| JP2020507106A (en) | Low resolution slide imaging, slide label imaging and high resolution slide imaging using dual optical paths and a single imaging sensor | |
| EP2828700A1 (en) | Multi-color confocal microscope and imaging methods | |
| JP2010080144A (en) | Compound microscope device and method of observing sample | |
| JP4725967B2 (en) | Minute height measuring device and displacement meter unit | |
| CN113916776A (en) | Axial positioning system and method based on virtual pinhole confocal technology | |
| EP2733514A1 (en) | Apparatus for structured illumination of a specimen | |
| US20190043687A1 (en) | System and method for axial scanning based on static phase masks | |
| US20210325655A1 (en) | Stepped biological chip and gene sequencing device for testing the same | |
| JP5190603B2 (en) | Optical microscope and observation method | |
| CN111521561A (en) | Multi-mode microscopic hyperspectral imager |
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 | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20220111 |
|
| RJ01 | Rejection of invention patent application after publication |