CN100483110C - Parallel column scanning spectrum type surface plasma resonant imaging method and apparatus - Google Patents
Parallel column scanning spectrum type surface plasma resonant imaging method and apparatus Download PDFInfo
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- CN100483110C CN100483110C CNB2005100362323A CN200510036232A CN100483110C CN 100483110 C CN100483110 C CN 100483110C CN B2005100362323 A CNB2005100362323 A CN B2005100362323A CN 200510036232 A CN200510036232 A CN 200510036232A CN 100483110 C CN100483110 C CN 100483110C
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Abstract
Its method includes the following steps: changing the beam emitted by broadband light source into a hairline beam; making said hairline beam be irradiated on SPR generation device; after the beam outcome from said device is dispersed, utilizing two-dimensional photoelectric detection device to make detection and obtain SPR spectrum information in a lighted line-type region of tested plane; translating said device and using said hairline beam to scan the whole tested plane to obtain SPR spectrum information of all the columns of whole tested plane; utilizing computer to make operation and processing to obtain refractive index information of whole tested plane in said device so as to form a two-dimensional image. Said method can be used for detecting various biological chips.
Description
Technical field:
The invention belongs to optical imagery detection technique field, particularly relate to the refractive index imaging method and apparatus of a kind of spectrum type surface plasma resonant (SPR) technology based on column scan.
Background technology:
Surface plasma body resonant vibration (surface plasmon resonance is abbreviated as SPR) method for sensing, highly sensitive and need not the characteristics of mark because of it, be widely used in the biological and chemical field.The SPR method for sensing of useful point measurement is studied the report (Anal.Chem.70 (1998) 2019-2024) of albumino reaction, chemical reaction etc.But along with the widespread use of biologic arrays (being biochip) such as gene array, protein arrays, the refractive index sensing of point more and more can not satisfy the demand of research, needs the method that can survey two dimensional surface---be imaging detects.And traditional formation method that utilizes SPR (Nature 332 1988.615-616.), be SPM (surface plasmon microscopy), the fixedly frequency of incident parallel beam and incident angle, directly reflected light is carried out imaging, the gradation of image value that is become not is a linear relationship with the corresponding of refractive index, so change of refractive can only be described qualitatively; And this method can only measure and change little ranges of indices of refraction, surpasses this scope, and two different refractive indexes may corresponding identical gray-scale values; Part is used this class device of laser lighting in addition, because the strong coherence of laser can produce and dodge the spot effect, has a strong impact on imaging effect.
So people begin to seek other and utilize the SPR phenomenon to come imaging method.People such as Knobloch use the chromatic dispersion SPM of white-light illuminating begun to utilize spectrum SPR imaging research (Appl.Phys.Lett.69,2336-23371996), but this method can only provide refractive index information qualitatively with common SPM is the same;
The a branch of very thin white light beam of humans such as Yuk is fixed into firing angle and is radiated on the SPR device, excites the SPR of a point on the tested surface, and reflected light obtains the resonant wavelength of this point with spectrometer analysis; Each point on the two-dimensional scan plane, the resonant wavelength that obtains whole plane distributes, (Proteomics 2004,4,3468-3476 thereby calculate the out of Memory such as index distribution on whole plane; Biosensors﹠amp; Bioelectronics 20 (11) 2189-2196,2005).This method can be described the index distribution of tested surface quantitatively, but this method adopts a point, a some ground scanning survey, and measuring speed is very slow like this, can not adapt to the requirement of the contour flux measurement of protein arrays.
Summary of the invention
In view of existing two-dimentional SPR imaging detection technology above shortcomings, the invention provides a kind of Parallel column scanning spectrum type surface plasma resonant imaging method and apparatus, this method is utilized spectral type SPR technology, adopt linear beam to carry out the column scan of one dimension, measure the index distribution image of two dimension.Utilize this method, can survey whole refractive index information of measured in high flux ground, the two dimensional image that obtains, its every bit gray-scale value are directly linearly corresponding to tested surface corresponding point refractive index.
Parallel column scanning spectrum type surface plasma resonant imaging method of the present invention comprises the steps:
A, the light beam that wideband light source is sent are transformed into a fine rule shaped light beam, described fine rule shaped light beam is radiated on the SPR generating means, after the outgoing beam of SPR generating means carries out chromatic dispersion via a dispersion means, by the 2 D photoelectric detection, obtain the illuminated interior row SPR spectrum picture information of a linear areas in tested plane in the SPR generating means, be transferred to computer storage then;
B, make tested plane in the described SPR generating means in this plane along direction translation perpendicular to the field of illumination, scan whole tested plane by described linear beam, and in this process, use 2 D photoelectric sniffer continuous probe, obtain whole row SPR spectrum picture information on whole tested plane;
C, computing machine carry out computing, processing to described whole row SPR spectrum picture information, obtain the refractive index information on whole tested plane in the described SPR generating means, and generate width of cloth two dimension index distribution picture.
Realize a kind of parallel column scanning spectrum type surface plasma resonant imaging device of above-mentioned formation method, mainly comprise: SPR generating means, incident arm, reflection arm and Flame Image Process computing machine; Described SPR generating means can translation, and it is installed on the controlled mobile platform; Described incident arm is arranged at a side of SPR generating means, and it comprises wideband light source, polarizer and produce the optical system that contains the cylindricality mirror of a fine rule shaped light beam that described fine rule shaped light beam is injected the tested plane of SPR generating means successively; Described reflection arm comprises dispersion means and the 2 D photoelectric sniffer that is used for chromatic dispersion successively; The output terminal of 2 D photoelectric sniffer is connected with the respective input of computing machine, and each row SPR spectrum picture information that the 2 D photoelectric sniffer detects all is transferred to computer storage, processing, and generates width of cloth two dimension index distribution picture.
The present invention is as the SPR imaging, and the refractive index with all SPR technology changes responsive characteristics, and higher refractive index resolution is arranged.Compared with prior art, the present invention also has the advantage of following uniqueness:
1, the inventive method imaging has reflected the index distribution on measured plane quantitatively, truly.
2, owing to adopt linear beam once to obtain one-dimension information, column scan obtains two-dimensional signal, and the present invention has the advantages that information throughput is big, efficient is high, can be widely used in biology, chemical field, is very suitable for the application in various biochips field.
3, just can obtain whole refractive index information of tested one dimension each point in the one-shot measurement of the present invention, therefore also can not carry out column scan, use as the one dimension spr sensor.
4, the present invention can come the light conducting spectrum information with optical fiber owing to adopt spectral measurement, and the SPR test surface can carry out the measurement of remote and particular surroundings (as poisonous, inflammable and explosive environment) away from light source and analytical equipment like this.
Description of drawings
Fig. 1 is the theory diagram of the parallel scan light spectral pattern SPR imaging device of the present invention;
Fig. 2 is whole row image information flow process figure on the whole tested plane of its COMPUTER CALCULATION, processing acquisition;
Fig. 3 is a row image information flow process figure on the whole tested plane of its COMPUTER CALCULATION, processing acquisition;
Fig. 4 is the example picture that adopts a glycerine above the golden film that the inventive method records;
Fig. 5 is an one embodiment device synoptic diagram.
Embodiment
The present invention is further described below in conjunction with accompanying drawing.
With reference to Fig. 1, the parallel column scanning spectrum type surface plasma resonant imaging device mainly comprises SPR generating means 4, incident arm, reflection arm and Flame Image Process computing machine 7.Described SPR generating means 4 comprises prism and the golden film that is arranged on the prism inclined-plane, and the outside surface of this gold film is tested plane, is used to adhere to by the side thing; Described SPR generating means 4 can translation, and it is installed on the controlled mobile platform 43, and this controlled mobile platform can adopt manually or automatic control mode is realized.Described incident arm is arranged at a side of SPR generating means 4, and it comprises wideband light source 1, polarizer 2 and produce the optical system that contains the cylindricality mirror 3 of a fine rule shaped light beam that described fine rule shaped light beam is injected the tested plane of SPR generating means 4 successively; Described reflection arm is arranged at the opposite side of SPR generating means 4, and it comprises dispersion means 5 and the 2 D photoelectric sniffer 6 that is used for chromatic dispersion successively; The output of 2 D photoelectric sniffer is connected with the input of computing machine 7, and each row image information that the 2 D photoelectric sniffer detects all is transferred to computing machine 7 storages, handles, and generates a width of cloth two-dimension picture.
The dispersion means 5 of reflection arm can comprise the lens 52,54 before and after slit 51, grating 53 and the grating, and dispersion means 5 also can adopt a grating monochromator to realize.The optical system that contains the cylindricality mirror 3 of incident arm can adopt a cylindrical convex lens or cylindrical mirror.Wideband light source 1 can adopt Halogen lamp LED or broadband light sources such as light emitting diode and combination or mercury lamp.
The present invention can come the light conducting spectrum information with optical fiber owing to adopt spectral measurement, and promptly the outgoing beam of the fine rule shaped light beam of SPR generating means 4 plane of incidence sides, SPR generating means 4 reflecting surface sides all comes conduct far by optical fiber.The SPR test surface can carry out the measurement of remote and particular surroundings (as poisonous, inflammable and explosive environment) away from light source and analytical equipment like this.
The light that wideband light source 1 sends becomes polarized light through polarizer 2 (as polaroid, polarizing prism etc.), becomes a fine rule shaped light beam through an optical system 3 that contains the cylindricality mirror again.This linear beam is incident on the SPR generating means 4, and the optical direction of polarizer 2 is adjusted into the p light direction of SPR device 4.The outgoing beam of SPR generating means 4, carry out chromatic dispersion via a dispersion means 5 after, detect by 2 D photoelectric sniffer 6, be transferred to computing machine 7 places imaging.A row image that obtains has like this comprised the spectral information of the SPR in the illuminated linear areas of tested plane in the SPR generating means 4.
Drive by controlled mobile platform 43, the tested plane that makes SPR generating means 4 in this plane along direction translation perpendicular to the field of illumination, scan whole plane with described linear beam, and it is continuous in 6 imagings of 2 D photoelectric sniffer in this process, obtain whole row image informations on whole tested plane, a series of images that obtains has so just comprised the SPR spectral information on whole tested plane.
With reference to Fig. 2, the process of whole row image informations on COMPUTER CALCULATION, the whole tested plane that handle to obtain can comprise: 1), scan tested plane with linear beam, use 2 D photoelectric sniffer continuous probe simultaneously, obtain a series of figure; 2), get a wherein width of cloth picture; 3), handle this width of cloth of row image information process analysis figure, obtain a row refractive index information according to Fig. 3; 4), write down this row refractive index information; 5), repeat above 2) to 4) step, until handling all pictures, obtain a two-dimensional array that comprises refractive index information like this; 6), the refractive index linearity is scaled gray-scale value, a width of cloth figure draws.The quantitative refractive index profile of Here it is lining face.
With reference to Fig. 3, the process of Computer Processing one row image information comprises: 1), a width of cloth figure that the 2 D photoelectric sniffer is obtained, and according to of the spectral response curve correction of 2 D photoelectric sniffer to different wave length; 2), get the data line of this image; 3), the minimum value respective number of pixels of this line data is tried to achieve in match; 4), try to achieve corresponding SPR resonant wavelength by pixel count; 5) and then by the SPR resonant wavelength try to achieve corresponding refractive index; 6), write down the refractive index of this row; 7), the next line data of getting this image, repeat above 2) to 6) step, finish until data processing to all row of picture, so just obtained a row refractive index information.
Fig. 5 represents a specific embodiment device.Wideband light source 1 adopts the white point light source, as Halogen lamp LED, and the photoelectricity GCO-13M of Daheng, 100W.The light that this white point light source sends, through an achromatic imaging len 21 (as f=50mm, the photoelectricity GCL-010630 of Daheng) becomes directional light, by the polarization direction polaroid 22 (as the photoelectricity GCL-050002 of Daheng) that is horizontal direction, through a cylindrical convex lens 3 (as the photoelectricity GCL-110101 of Daheng), it is assembled in the horizontal direction become a fine rule shaped light beam again.When illumination is to SPR generating means 4 like this, is similar to and has formed a line spot.
SPR generating means 4 adopts Kretschmann total reflection model (Z.Naturforsch.23A (1968) 2135-2136.): plated the thick golden film 42 of 45nm on the inclined-plane of BK7 prism 41, golden film is tested plane outward.SPR generating means 4 is installed in a controlled one dimension mobile platform 43, can allow prism above it along the direction as shown translation.Above-mentioned fine rule shaped light beam is focused on the golden film, and the incident angle of central light beam is about 45 °.Apart from launching spot 10cm place, place the slit 51 of a 0.1mm, in the SPR reflected light, pick out the reflected light of a certain reflection angle.Light through slit becomes directional light by an imaging len 52, carries out chromatic dispersion through a reflection grating 53, through an imaging len 54, be imaged on the CCD device 6, the CCD device can be Q-imaging RETIGA EXi again, 12,1392 * 1040 pixels.Lens the 52, the 54th in the example before and after slit 51, grating 53 and the grating are realized by using a grating monochromator, as: ACTON RESEARCHSpectraPro 150, grating 300 lines.
Each row of resulting like this image after the response curve correction of CCD device to the light of different wave length, is exactly the SPR spectral distribution of the corresponding point of the linear field of illumination on the golden film.We can find the wavelength of reflectivity minimum---and the SPR resonant wavelength again by the corresponding relation of SPR resonant wavelength and refractive index, can obtain the refractive index of this point.Therefore this width of cloth figure has comprised the refractive index information of whole points of linear areas.Translation SPR plane with the tested plane of linear beam scanning survey, so just can obtain the refractive index information on whole plane.Generate a width of cloth figure,, go up every bit for figure and give one and put the gray-scale value that corresponding refractive index becomes linear dependence, just obtained the refractive index profile on tested plane with this according to the distribution range of refractive index in the tested surface.As Fig. 4 is the example picture that adopts a glycerine above the golden film that the inventive method records.
Claims (8)
1, a kind of parallel column scanning spectrum type surface plasma resonant imaging method is characterized in that comprising the steps:
A, the light beam that wideband light source is sent are transformed into a fine rule shaped light beam, described fine rule shaped light beam is radiated on the SPR generating means, after the outgoing beam of this device carries out chromatic dispersion via a dispersion means, by the 2 D photoelectric detection, obtain the illuminated interior row SPR spectrum picture information of a linear areas in tested plane in this device, be transferred to computer storage then;
B, make tested plane in the described SPR generating means in this plane along direction translation perpendicular to the field of illumination, scan whole tested plane by described linear beam, and in this process, use 2 D photoelectric sniffer continuous probe, obtain whole row SPR spectrum picture information on whole tested plane;
C, computing machine carry out computing, processing to described whole row SPR spectrum picture information, obtain the refractive index information on whole tested plane in the described SPR generating means, and generate width of cloth two dimension index distribution picture;
Wherein, the process of whole row SPR spectrum picture information on COMPUTER CALCULATION, the described whole tested plane of processing may further comprise the steps:
1), gets a wherein width of cloth picture;
2), according to handling this width of cloth of row image information process analysis figure, obtain a row refractive index information, specifically may further comprise the steps:
2.1), a width of cloth figure that the 2 D photoelectric sniffer is obtained, according to of the spectral response curve correction of 2 D photoelectric sniffer to different wave length;
2.2), get the data line of this image;
2.3), match tries to achieve the minimum value respective number of pixels of this line data;
2.4), try to achieve corresponding SPR resonant wavelength by pixel count;
2.5) and then try to achieve corresponding refractive index by the SPR resonant wavelength;
2.6), write down the refractive index of this row;
2.7), the next line data of getting this image, repeat above 2.2) to 2.6) step, finish until data processing to all row of picture, so just obtained a row refractive index information;
3), write down this row refractive index information;
4), repeat above 1) to 3) step, until handling all pictures, obtain a two-dimensional array that comprises refractive index information like this;
5), the refractive index linearity is scaled gray-scale value, obtain the quantitative refractive index profile of lining face.
2, according to the parallel column scanning spectrum type surface plasma resonant imaging method of claim 1, it is characterized in that: in step a, the light that described wideband light source sends is transformed into polarized light by polarizer, becomes a fine rule shaped light beam through an optical system that contains the cylindricality mirror again.
3, according to the parallel column scanning spectrum type surface plasma resonant imaging method of claim 1, it is characterized in that: in step b, described SPR generating means is by a controlled mobile platform translation, and described controlled mobile platform adopts manually or automatic control mode.
4, according to the parallel column scanning spectrum type surface plasma resonant imaging method of claim 1, it is characterized in that: the outgoing beam of the fine rule shaped light beam of described SPR generating means plane of incidence side, SPR generating means reflecting surface side is all by the optical fiber conduct far.
5, parallel column scanning spectrum type surface plasma resonant imaging device comprises SPR generating means (4), incident arm, reflection arm and Flame Image Process computing machine (7), it is characterized in that:
Described SPR generating means (4) can translation, and it is installed on the controlled mobile platform (43);
Described incident arm is arranged at a side of SPR generating means (4), it comprises wideband light source (1), polarizer (2) and produces the optical system that contains the cylindricality mirror (3) of a fine rule shaped light beam that described fine rule shaped light beam is injected the tested plane of SPR generating means (4) successively;
Described reflection arm comprises dispersion means (5) and the 2 D photoelectric sniffer (6) that is used for chromatic dispersion successively; The output terminal of 2 D photoelectric sniffer is connected with the respective input of computing machine (7);
Image processing software is installed in the described computing machine (7), each row SPR spectrum picture information that the 2 D photoelectric sniffer detects all be transferred to computing machine (7) storage and by image processing software by after the following step process, generate width of cloth two dimension index distribution picture;
1), gets a wherein width of cloth picture;
2), according to handling this width of cloth of row image information process analysis figure, obtain a row refractive index information, specifically may further comprise the steps:
2.1), a width of cloth figure that the 2 D photoelectric sniffer is obtained, according to of the spectral response curve correction of 2 D photoelectric sniffer to different wave length;
2.2), get the data line of this image;
2.3), match tries to achieve the minimum value respective number of pixels of this line data;
2.4), try to achieve corresponding SPR resonant wavelength by pixel count;
2.5) and then try to achieve corresponding refractive index by the SPR resonant wavelength;
2.6), write down the refractive index of this row;
2.7), the next line data of getting this image, repeat above 2.2) to 2.6) step, finish until data processing to all row of picture, so just obtained a row refractive index information;
3), write down this row refractive index information;
4), repeat above 1) to 3) step, until handling all pictures, obtain a two-dimensional array that comprises refractive index information like this;
5), the refractive index linearity is scaled gray-scale value, obtain the quantitative refractive index profile of lining face.
6, according to the parallel column scanning spectrum type surface plasma resonant imaging device of claim 5, it is characterized in that: the described optical system (3) that contains the cylindricality mirror adopts a cylindrical convex lens or cylindrical mirror.
7, according to the parallel column scanning spectrum type surface plasma resonant imaging device of claim 5, it is characterized in that: the dispersion means of described reflection arm (5) comprises that lens (52,54) or the dispersion means (5) before and after slit (51), grating (53) and the grating adopts a grating monochromator.
8, according to the parallel column scanning spectrum type surface plasma resonant imaging device of claim 5, it is characterized in that: described wideband light source (1) adopts Halogen lamp LED or light emitting diode or mercury lamp.
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| CN106198453B (en) * | 2016-08-25 | 2019-03-26 | 清华大学深圳研究生院 | A kind of online test method and device of the chief and vice reaction ratio of all-vanadium flow battery |
| CN107064106B (en) * | 2017-04-11 | 2019-07-05 | 北京航空航天大学 | A kind of two-dimensional surface plasma most preferably excites the high-precision recognition methods of Angle Position |
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| Sensitivity of ex situ and in situspectralsurfaceplasmonresonance sensors in the analysis ofproteinarrays. Jong Seol Yuk, Jae-Wan Jung, Se-HuiJung,Jeong-AHan,Young-Myeong Kim and Kwon-Soo Ha.Biosensors and Bioelectronics,Vol.20 . 2005 |
| Sensitivity of ex situ and in situspectralsurfaceplasmonresonance sensors in the analysis ofproteinarrays. Jong Seol Yuk, Jae-Wan Jung, Se-HuiJung,Jeong-AHan,Young-Myeong Kim and Kwon-Soo Ha.Biosensors and Bioelectronics,Vol.20 . 2005 * |
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