CN110672883B - Near-field super-resolution optical imaging method based on periodic nanopore array and lens medium microsphere array - Google Patents
Near-field super-resolution optical imaging method based on periodic nanopore array and lens medium microsphere array Download PDFInfo
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- CN110672883B CN110672883B CN201910987418.9A CN201910987418A CN110672883B CN 110672883 B CN110672883 B CN 110672883B CN 201910987418 A CN201910987418 A CN 201910987418A CN 110672883 B CN110672883 B CN 110672883B
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01Q—SCANNING-PROBE TECHNIQUES OR APPARATUS; APPLICATIONS OF SCANNING-PROBE TECHNIQUES, e.g. SCANNING PROBE MICROSCOPY [SPM]
- G01Q60/00—Particular types of SPM [Scanning Probe Microscopy] or microscopes; Essential components thereof
- G01Q60/18—SNOM [Scanning Near-Field Optical Microscopy] or apparatus therefor, e.g. SNOM probes
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- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B27/00—Optical systems or apparatus not provided for by any of the groups G02B1/00 - G02B26/00, G02B30/00
- G02B27/58—Optics for apodization or superresolution; Optical synthetic aperture systems
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- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B5/00—Optical elements other than lenses
- G02B5/008—Surface plasmon devices
Abstract
The invention discloses a near-field super-resolution optical imaging method based on a periodic nanopore array and a lens medium microsphere array, wherein incident light can be split into fine light beams with the diameter of 10-500nm through the periodic nanopore array, and the light beams can be respectively focused by the lens medium microsphere array to form a light spot array. The hole array corresponds to the central shaft of the lens medium microsphere array through the three-dimensional moving period nanopore array device, and light beams dispersed by the hole array are focused to the maximum extent to obtain a nanoscale light spot array, so that the purpose of super-resolution microscopic imaging is achieved. The invention has the advantages that the advantages of the periodic nanopore array and the lens medium microsphere array are combined, the point scanning mode of the traditional approach scanning optical microscope is expanded into array scanning, the defects that the single imaging range is small, an ultra-fine probe is needed, the scanning process is contacted with a sample, and the sample with large height change cannot be imaged are overcome, and the ultrahigh-resolution microscopic imaging is realized.
Description
Technical Field
The invention relates to the technical field of near-field super-resolution optical imaging, in particular to a near-field super-resolution optical imaging method based on a periodic nanopore array and a lens medium microsphere array.
Background
In 1928, england scientist Synge proposed for the first time an assumption of being able to break through the resolution of an optical microscope, which assumed that a sub-wavelength detector was placed in the Near-field range of a sample, and Scanning and collecting the optical signal of each point by point, so as to realize super-resolution imaging, namely Scanning Near-field optical microscope (SNOM) (Synge E H.A shielded method for extended micro-optical resolution in the ultra-micro-optical region. phosphor optical microscope, 1928,6 (356): 362). Synge describes this method in detail by placing an opaque plate with a small diameter aperture at a distance of less than 100nm from the sample surface, and passing the incident light through the aperture to become a very small spot. Two-dimensional imaging of the sample can be obtained by illuminating the sample with small spots, scanning point by point in steps of 10nm, and detecting the light from the sample, which technique, according to his prophetic, can increase the optical resolution to the nanometer level. The key point of the SNOM capable of realizing super-resolution imaging is that 1) the light source is strong enough; 2) a subwavelength-scale aperture; 3) nano-scale movement and positioning accuracy.
In the near-field optical region, the size of the probe determines the resolution of the probe, regardless of the wavelength of the illumination source (wang, wudayu, sun lin. However, the aperture probe has low laser coupling efficiency and weak signal. When the aperture diameter reaches 100nm, the light transmittance sharply decreases. There is therefore a need to select and balance between optical resolution and luminous efficiency. In practical application, the SNOM resolution of the current aperture probe does not exceed 50 nm. Meanwhile, the practical manufacturing of the nano probe is limited by various factors and various process conditions, and is different from the ideal design. If the tip of the needle tip is too large and too thick; the aperture is not a circular hole; the probe is contaminated; the metal film has defects and even partially warps; the film coating damages aperture light leakage and the like, which can seriously affect the SNOM imaging quality. The SNOM development suffers from the additional limiting factor of lower temporal resolution. Longer scan times are required for large area and high resolution imaging and studies of single molecule localization and dynamic processes in cells are not satisfactory.
Disclosure of Invention
The invention provides a super-resolution optical microscopic imaging method based on a periodic nanopore array and a lens medium microsphere array, aiming at the problems of low resolution and limited imaging speed of the existing near-field scanning optical microscope. Compared with a near-field scanning optical microscope which scans point by point, the scheme has higher imaging speed and can improve the spatial resolution of the microscope.
In order to achieve the purpose, the technical scheme provided by the invention is as follows:
a near-field super-resolution optical imaging method based on a periodic nanopore array and a lens medium microsphere array comprises the following steps:
s1, adding a three-dimensional piezoelectric displacement table above the near-field optical microscope, wherein the three-dimensional piezoelectric displacement table is sequentially provided with a focusing module consisting of an optical fiber, a collimator, a periodic nanopore array and a lens medium microsphere array from top to bottom, the hole pitch of the periodic nanopore array is the same as the microsphere pitch in the lens medium microsphere array, and holes are kept to correspond to microsphere central axes;
s2, placing the sample on the focal plane of the lens medium microsphere array and within the imaging range of the near-field microscope;
s3, completing the image acquisition work of the sample in the visual field in a scanning dot array mode;
s4, controlling the focusing module to gradually and sequentially move on the surface of the sample by using the three-dimensional piezoelectric displacement table until the image acquisition of the surface of the sample to be detected is completed;
and S5, splicing all the collected images in sequence to realize the super-resolution image of the surface of the sample to be detected.
Preferably, the lens medium microsphere array is a transparent silica microsphere array, a polystyrene microsphere array or a quartz microsphere array.
Preferably, the microsphere size in the lens medium microsphere array is 500nm-10 μm.
Preferably, the pore size of the periodic nanopore array is 10-500nm, and the pore shape is circular, square or regular polygon.
Preferably, the thickness of the metal light-shielding film on the periodic nanopore array is subject to no more than 5% of the light transmittance of incident light introduced by the optical fiber.
Preferably, the metal light shielding film on the periodic nanopore array is made of metals including gold, platinum, chromium and gold-palladium alloy.
Preferably, the wavelength of the incident light introduced by the optical fiber is in the range of 400nm to 1500 nm.
Preferably, the lens medium microsphere array is formed by self-assembling microspheres to form a membrane structure or fixing lens medium microspheres in a periodic frame structure to form an array.
Preferably, the preparation method of the periodic nanopore array comprises the steps of substrate cleaning, glue coating, electron beam exposure, top glue development, primer etching, photomask plating and glue washing.
Compared with the prior art, the invention has the technical effects that:
the near-field super-resolution optical imaging method based on the periodic nanopore array and the lens medium microsphere array adopts a method of focusing a light beam by the lens medium microsphere array, incident light is divided into very thin light beams through the periodic nanopore array, the light beams enter the lens medium microsphere array and are further focused into high-energy light spots, the SNOM signal is enhanced, the problem that the SNOM signal of a pore diameter probe is weak is solved, and meanwhile, the problem of low SNOM time resolution can be solved by upgrading point scanning to dot scanning. By designing the parameters of the nanopore periodic array, local field enhancement is realized by utilizing the characteristics of Surface Plasmon Polaritons (SPPs), and signal detection of the nanopore array is realized by selecting a single photon photoelectric detection array with high quantum efficiency. The invention relates to a near-field super-resolution optical microscope based on a periodic nanopore array and a lens medium microsphere array, which utilizes an optical fiber, a collimator, the periodic nanopore array and the lens medium microsphere array as focusing modules, improves the resolution of the near-field microscope, and realizes spatial super-resolution imaging. The point scanning can be upgraded to array scanning by using the focusing module, and compared with a near-field scanning optical microscope, the near-field scanning optical microscope has the advantages of higher imaging speed and larger imaging range.
Drawings
In order to more clearly illustrate the embodiments of the present application or technical solutions in the prior art, the drawings needed to be used in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments described in the present invention, and other drawings can be obtained by those skilled in the art according to the drawings.
Fig. 1 is a schematic structural diagram of a near-field super-resolution optical imaging device based on a periodic nanopore array and a lens medium microsphere array provided by the invention.
FIG. 2 is a flow chart of the periodic nanopore array fabrication of the present invention;
FIG. 3 is an atomic force microscope of a periodic nanopore array of the present invention;
FIG. 4 is an electron microscope image of a periodic nanopore array of the present invention;
FIG. 5 is an optical inspection of a periodic nanopore array of the present invention;
FIG. 6 is an optical inspection of a microsphere array of lens media according to the present invention.
Description of reference numerals:
1. an optical fiber; 2. a three-dimensional piezoelectric displacement stage; 3. a collimator; 4. a periodic array of nanopores; 5. a lens dielectric microsphere array; 6. a sample; 7. an objective lens; 8. an optical filter; 9. a tube mirror; 10. a detector.
Detailed Description
In order to make the technical solutions of the present invention better understood, those skilled in the art will now describe the present invention in further detail with reference to the accompanying drawings.
The invention provides a near-field super-resolution optical imaging method based on a periodic nanopore array and a lens medium microsphere array, which comprises the following steps of:
step S1, adding a three-dimensional piezoelectric displacement platform 2 above the near-field optical microscope, wherein the three-dimensional piezoelectric displacement platform 2 is sequentially provided with a focusing module consisting of an optical fiber 1, a collimator 3, a periodic nanopore array 4 and a lens medium microsphere array 5 from top to bottom, the hole pitch of the periodic nanopore array is the same as the microsphere pitch in the lens medium microsphere array, and holes are kept to correspond to the central axis of microspheres; the device structure is shown in fig. 1, and comprises a focusing module consisting of an optical fiber 1, a collimator 3, a periodic nanopore array 4 and a lens medium microsphere array 5, and an imaging system consisting of a sample 6, an objective lens 7, an optical filter 8, a tube mirror 9 and a detector 10.
The periodic nanopore array is a metal pore which is prepared on a glass substrate, has the aperture of 10-500nm and the spacing matched with the lens medium microsphere array, the preparation flow chart is shown in figure 2, and the preparation method comprises the following steps:
1) cleaning a substrate: the glass substrate is treated in a concentrated sulfuric acid: soaking in a solution of hydrogen peroxide 3:1 for 1 hour, washing with water and ethanol respectively, placing in a dryer for 2 hours, and cleaning with oxygen plasma for 3 minutes before use. Platinum is plated on a clean glass substrate as a conductive layer.
2) Gluing: coating a bottom layer of photoresist (PMMA 495k) by using a spin coater at the rotation speed of 1000-5000rpm, drying the photoresist in a glove box by using a heating plate, coating a top adhesive (HSQ-006) with the thickness of 30-60nm, and drying the photoresist again for later use.
3) Electron beam exposure: the appropriate electron beam dose (0.4-40mC/cm2), exposure voltage (10-100keV), current (1-500pA), and exposure time (1-1000s) were selected for precise exposure on the topcoat depending on the designed array pattern.
4) Gum-jacking development: and (4) selecting a developing solution aiming at the HSQ to develop the top glue to obtain the accurate hole array reverse template.
5) Etching the primer: and taking the top glue pattern as a mask plate, etching the bottom glue in a large range by using oxygen plasma, and transferring the pattern on the top glue to the bottom glue.
6) Plating a light shielding film: and plating a metal shading film on the etched substrate to ensure that the shading efficiency is over 95 percent.
7) Washing glue: and soaking the substrate with a photoresist removing solution aiming at the Primer (PMMA) to obtain the designed metal hole array.
The size of the microspheres in the lens medium microsphere array is 500nm-10 mu m, and the preparation method of the self-assembled film comprises the following steps:
1) and (5) cleaning the slide. The slide is soaked in acetone solution and ultrasonic treatment is carried out for 10 min.
2) And (4) carrying out double hydrophilic treatment on the slide. Moving the glass slide into a mixed solution of a sulfuric acid solution and hydrogen peroxide in a volume ratio of 3:1, soaking for 1h, transferring the glass slide out of the mixed solution, and cleaning the glass slide by using ultrapure water to remove residual substances;
3) the washed slide glass was silanized with 3-Aminopropyltriethoxysilane (APTES). The specific method comprises the following steps: and (3) placing the treated glass slide and a small container in a vacuum dryer, introducing argon for 5min, adding APTES into the small container, continuously introducing argon for 5min, and sealing the dryer for 4 h.
4) Diluting the microsphere solution in an ethanol solution according to the volume ratio of 1:2, and performing ultrasonic treatment in an ultrasonic instrument for 10min to obtain a mixed solution;
5) respectively dripping 200 μ L PS bead solution into water, standing for more than 12h, adding surfactant sodium dodecyl sulfate to promote film formation, inserting the slide modified by APTES at 45 deg.C below the liquid surface, and pulling out to form single layer film.
6) And (5) maintaining the temperature in an oven at 85 ℃ and drying.
And S2, placing the cell sample to be detected on the focal plane of the lens medium microsphere array and within the imaging range of the near-field microscope.
And step S3, finishing the acquisition work of the sample image in the visual field in a dot array scanning mode. Specifically, incident light guided by the optical fiber is parallelly incident into the periodic nanopore array after passing through the collimator, is split into small light beams, is focused to obtain light spots after passing through the lens medium microsphere array and is irradiated onto the surface of a sample, and is finally received by a detector through optical devices such as an objective lens, an optical filter and a tube lens, so that the acquisition of sample information is completed. The wavelength range of the incident light is 400nm-1500 nm.
And step S4, controlling the focusing module to gradually and sequentially move on the surface of the sample by using the three-dimensional piezoelectric displacement table until the image acquisition of the surface of the sample to be detected is completed.
And S5, splicing all the collected images in sequence to realize the super-resolution image of the surface of the sample to be detected. Atomic force microscopy, electron microscopy imaging and optical detection images of periodic nanopore arrays are shown in figures 3-5: the atomic force microscope image represents the specific characteristics of the periodic nanopore array such as appearance, size, hole spacing and the like, and proves that the preparation method can realize the preparation of the periodic nanopore array; the electron microscope further verifies the feasibility of the method; optical detection proves the light transmittance of the periodic nanopore array, and lays a foundation for the next step of coaxiality and focusing with the lens medium microsphere array. The optical detection graph of the lens medium microsphere array is shown in fig. 6, the feasibility of the preparation method of the lens medium microsphere self-assembled film can be verified through characterization of the lens medium microsphere self-assembled film, the self-assembled film is large in area and stable in structure, the number of microspheres in a unit area is large, and the light beams transmitted by the periodic nanopore array can be conveniently focused.
The traditional approach scanning optical microscope only has one probe, the scanning range is limited, the invention expands the point scanning into the array scanning, which is equivalent to the simultaneous imaging of a plurality of probes, and the imaging speed is accelerated by the high-flux imaging mode; in addition, a smaller light spot can be obtained through a focusing mode of combining the periodic nanopore array and the lens medium microsphere array, and the spatial resolution of the microscope is improved. In conclusion, the periodic nanopore array can split incident light into small light beams with the diameter of 10-500nm, and the light beams can be respectively focused through the microsphere array to form a light spot array. The hole array corresponds to the central shaft of the lens medium microsphere array through the three-dimensional moving period nanopore array device, and light beams dispersed by the hole array are focused to the maximum extent to obtain a nanoscale light spot array, so that the purpose of super-resolution microscopic imaging is achieved. The invention has the advantages that the advantages of the periodic nanopore array and the lens medium microsphere array are combined, the point scanning is expanded into array scanning, the defects that the near-field scanning optical microscope has a small imaging range, needs an extremely fine probe, is contacted with a sample in the scanning process and cannot image the sample with large height change are overcome, and the ultrahigh-resolution microscopic imaging is realized.
While certain exemplary embodiments of the present invention have been described above by way of illustration only, it will be apparent to those of ordinary skill in the art that the described embodiments may be modified in various different ways without departing from the spirit and scope of the invention. Accordingly, the drawings and description are illustrative in nature and should not be construed as limiting the scope of the invention.
Claims (6)
1. A near-field super-resolution optical imaging method based on a periodic nanopore array and a lens medium microsphere array is characterized by comprising the following steps:
s1, adding a three-dimensional piezoelectric displacement table above the near-field optical microscope, wherein the three-dimensional piezoelectric displacement table is sequentially provided with a focusing module consisting of an optical fiber, a collimator, a periodic nanopore array and a lens medium microsphere array from top to bottom, the hole pitch of the periodic nanopore array is the same as the microsphere pitch in the lens medium microsphere array, and holes are kept to correspond to microsphere central axes; the wavelength range of the incident light guided by the optical fiber is 400nm-1500 nm; the aperture size of the periodic nanopore array is 10-500nm, and the thickness of the metal shading film on the periodic nanopore array is determined by that the light transmittance of incident light led in by the optical fiber is not more than 5%; the size of the microspheres in the lens medium microsphere array is 500nm-10 mu m;
s2, placing the sample on the focal plane of the lens medium microsphere array and within the imaging range of the near-field microscope;
s3, completing the image acquisition work of the sample in the visual field in a scanning dot array mode;
s4, controlling the focusing module to gradually and sequentially move on the surface of the sample by using the three-dimensional piezoelectric displacement table until the image acquisition of the surface of the sample to be detected is completed;
and S5, splicing all the collected images in sequence to realize the super-resolution image of the surface of the sample to be detected.
2. The near-field super-resolution optical imaging method based on the periodic nanopore array and the lens dielectric microsphere array according to claim 1, wherein the lens dielectric microsphere array is a transparent silica microsphere array, a polystyrene microsphere array or a quartz microsphere array.
3. The near-field super-resolution optical imaging method based on the periodic nanopore array and the lens medium microsphere array according to claim 1, wherein the shape of the pores of the periodic nanopore array is circular or regular polygon.
4. The near-field super-resolution optical imaging method based on the periodic nanopore array and the lens medium microsphere array of claim 1, wherein the metal light shielding film on the periodic nanopore array is made of metals including gold, platinum, chromium and gold-palladium alloy.
5. The near-field super-resolution optical imaging method based on the periodic nanopore array and the lens medium microsphere array according to claim 1, wherein the lens medium microsphere array is formed by self-assembling microspheres to form a membrane structure or fixing lens medium microspheres in a periodic frame structure.
6. The near-field super-resolution optical imaging method based on the periodic nanopore array and the lens medium microsphere array according to claim 1, wherein the preparation method of the periodic nanopore array comprises the steps of substrate cleaning, glue coating, electron beam exposure, top glue development, primer etching, shading film plating and glue washing.
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