CN110095441B - Fluorescent nanometer scale component and preparation and application thereof - Google Patents
Fluorescent nanometer scale component and preparation and application thereof Download PDFInfo
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- CN110095441B CN110095441B CN201910318120.9A CN201910318120A CN110095441B CN 110095441 B CN110095441 B CN 110095441B CN 201910318120 A CN201910318120 A CN 201910318120A CN 110095441 B CN110095441 B CN 110095441B
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- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
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Abstract
The invention relates to the technical field of ultrahigh resolution fluorescence microscopic imaging, in particular to a fluorescent nanometer scale component and preparation and application thereof; the fluorescent nanometer scale component comprises a substrate layer and a mask layer, grooves are arranged at intervals on the corresponding positions of the mask layer, the cross section width of each groove is 10-200nm, and the size (width) of each mask groove is small, so that the fluorescent nanometer scale assembled by the fluorescent nanometer scale component is accurate, and the requirement of calibrating and measuring the resolution of an ultrahigh-resolution fluorescence microscope can be met.
Description
Technical Field
The invention relates to the technical field of ultrahigh resolution fluorescence microscopic imaging, in particular to a fluorescent nanometer scale component and preparation and application thereof.
Background
Fluorescence microscopy imaging technology plays an important role in research in the field of life science, and can research the life process and mechanism in cells at the sub-cellular level, however, due to the limitation of Abbe diffraction limit in traditional optics, the imaging resolution cannot break through below 200 nm. With the rise of ultra-high resolution fluorescence microscopy, such as light-sensitive positioning microscopy (PALM), fluorescence light-sensitive positioning microscopy (FPALM), random optical reconstruction microscopy (STORM), etc., stimulated emission depletion microscopy (STED), structured light microscopy (SIM), single molecule positioning microscopy, etc., these techniques enable a fluorescence microscope to observe extremely fine structures of 20nm to 100 nm.
With the continuous progress and commercial promotion of the ultra-high resolution fluorescence microscopy, more and more universities, research institutes and companies begin to independently develop or directly purchase ultra-high resolution fluorescence microscopy instruments for life science research. During the daily use and maintenance of the instrument, calibration measurements are often required for the resolution of the fluorescence microscope, and therefore a standard structure of fluorescence with defined dimensions is used. To this end, chinese patent document CN103712965A first proposes an equidistant fluorescent nano standard plate based on nano-groove filled fluorescent quantum dots, and although the equidistant structure form of the plate meets the requirement of resolution determination, the processing method of the plate is complicated, and if the first method introduced therein has too many preparation steps, especially the prepared nano line structure cannot be formed at one time, and the quantum dot nano line structure with a certain aspect ratio can be obtained by assembling the quantum dot nano line structure for many times, the preparation steps are as follows: the method comprises the steps of firstly, spin-coating a photoresist layer 104 on a substrate 100, secondly, forming photoresist line structures with equal intervals through electron beam lithography to expose the substrate 100, thirdly, adsorbing a charged polyelectrolyte polymer on the exposed substrate 100 to form a layer 109, fourthly, forming a multilayer quantum dot nano-line structure 107 on the surface of the layer 109 through a Laber-by-layber assembly technology, wherein the quantum dot nano-line structure 107 needs to be arranged between 3 and 10 layers, fifthly, removing the photoresist line structures, and sixthly, spin-coating a polymer layer or other protective layers 108 on the surface of the layer 107.
In order to solve the problem of complicated preparation steps of the equidistant fluorescent nano standard plate, chinese patent document CN103954600A proposes a wide-line narrow-interval fluorescent nano scale structure based on fluorescent dye filled in a groove, but the wide-line narrow-interval fluorescent nano scale structure is wideThe structure form of narrow line interval does not meet the determination standard of resolution, can only be used for general debugging observation of fluorescence microscope, and it adopts photoresist as mask, because general photoresist can not reach such high aspect ratio as 30nm width, 100nm thickness, the slot size that causes to be prepared is limited, in order to solve the above-mentioned problem, in its preparation scheme, choose to keep the non-transparent conducting layer on the substrate first, then plate a layer of Si with strong anti-etching ability on it3N4Then spin-coating photoresist, exposing according to predetermined pattern to dissolve the photoresist and sequentially carrying out Si etching3N4The layer and the non-transparent conducting layer are etched, however, even if the groove structure with smaller size and larger depth-to-width ratio is prepared by selecting the scheme, the scale structure is larger in size, the scale is not accurate, the mask layer is small in thickness and poor in light blocking effect, and the non-transparent conducting layer below the mask layer is lower in melting point like an aluminum film, so that the scale structure is unstable under the condition of high-energy laser, easy to deform and short in service life.
Disclosure of Invention
Therefore, the technical problem to be solved by the present invention is to overcome the defect of inaccuracy of the fluorescent nano-scale component in the prior art, and further to provide a high-precision fluorescent nano-scale component, and a method and an application for preparing the fluorescent nano-scale component disclosed by the present invention.
To this end, the invention provides a fluorescent nanoscaler component comprising a substrate and a mask layer formed on the substrate, the mask layer comprising spaced apart grooves having a cross-sectional width of 10-200 nm.
Preferably, the grooves are arranged at equal intervals, and the width of the cross section of each groove is 10-150 nm; preferably, the depth-to-width ratio of the groove is 0.1-20.
Preferably, the material of the mask layer is selected from a high melting point metal or a non-metal material that is not transmissive to ultraviolet and visible light.
Preferably, the refractory metal is selected from one of Ta, Mo, Cr, Ti and Pd; the non-metallic material which can not transmit ultraviolet and visible light is Si or high-density carbon-doped polymer.
Preferably, when the material of the mask layer is high-melting-point metal, the thickness of the mask layer is 20-100 nm; when the material of the mask layer is Si, the thickness of the mask layer is 100-200 nm.
The invention also discloses a preparation method of the fluorescent nanometer scale component, which comprises the following steps:
treating the substrate: cleaning the surface of the substrate;
mask layer: formed on the surface of the substrate;
coating a photoresist: coating photoresist on the mask layer to form a photoresist mold layer with a thickness of 20-500 nm;
photoetching: patterning the photoresist mold layer, and forming photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of each groove is 10-200 nm;
transferring the pattern: transferring the photoetching pattern of the photoresist mould layer to the mask layer, and then removing the photoresist.
The invention also discloses a preparation method of the fluorescent nanometer scale component, which comprises the following steps:
treating the substrate: cleaning the surface of the substrate;
coating a photoresist: coating photoresist on a substrate to form a photoresist mold layer with the thickness of 20-500 nm;
photoetching: patterning the photoresist mold layer, and forming photoetching patterns of photoresist nano line structures arranged at intervals on the substrate, wherein the transverse cross section width of each photoresist nano line structure is 10-200 nm;
transferring the pattern: and depositing a mask layer material on one side of the substrate with the photoetching pattern to form a mask layer, then removing the photoresist, and transferring the photoetching pattern onto the mask layer.
Preferably, the photoresist layer is patterned by electron beam lithography, and the voltage of the electron beam lithography is 30kV-100 kV.
Preferably, when the high melting point metal is used as the mask layer material, the high melting point metal is deposited on the substrate by deposition, sputtering or electron beam evaporation, and the deposition speed is 0.1-0.5 nm/s.
Preferably, when Si is used as the mask layer material, the SOI substrate is subjected to HF etching and Si film aqueous phase transfer before the photoresist is applied, and Si is attached to the substrate as the mask layer material.
Preferably, in the transferring the pattern step, the photoresist pattern is transferred onto the mask layer using RIE or wet etching.
Preferably, the wet etching step comprises a gold-assisted etching step.
Preferably, before the photolithography step, a conductive polymer layer is further coated on the photoresist or a conductive metal layer is deposited on the photoresist.
The invention also discloses an application of the fluorescent nano scale component or the fluorescent nano scale component prepared by the preparation method in the field of microscopic fluorescence analysis.
The technical scheme of the invention has the following advantages:
1. the fluorescent nano scale component comprises a substrate and a mask layer formed on the substrate, grooves are arranged at intervals on the corresponding positions of the mask layer, the cross section width of each groove is 10-200nm, and the size (width) of each mask groove is small, so that the fluorescent nano scale assembled by the fluorescent nano scale component is accurate, and the requirement of calibrating and measuring the resolution of an ultrahigh-resolution fluorescence microscope can be met.
2. According to the fluorescent nanometer scale component, the grooves are arranged at equal intervals, and the width of the cross section of each groove is 10-150 nm; the grooves are arranged at equal intervals, so that the defect that the resolution measurement standard of a microscope is not met due to the structural form of wide lines and narrow intervals in the conventional fluorescent nanometer scale component is overcome, and the requirement of calibrating and measuring the resolution of the ultrahigh-resolution fluorescent microscope can be met; the width of the cross section of the groove is 10-150nm, so that the accuracy of the fluorescent nanometer scale is further improved;
furthermore, the depth-to-width ratio of the groove is 0.1-20, the thickness of the groove mask is further controlled to be 20-200nm, the depth-to-width ratio can reach 200nm/10nm at most, the light blocking effect is good, and the ruler precision is high.
3. The mask layer material of the fluorescent nanometer scale component is selected from high-melting-point metal, Si and high-density carbon-doped polymers, and has better stability when in use, and the mask layer material cannot deform due to long-time irradiation of high-energy laser, so that the service life is prolonged; because the high-melting-point metal is a conductor, the Si and high-density carbon-doped polymer belongs to a semiconductor, a conductive material layer is not needed in the process, only a substrate and a mask layer are needed, and the process is simple.
4. The preparation method of the fluorescent nanometer scale component adopts an electron beam lithography mode, a high-melting-point metal mask material or a Si film is attached to a substrate before photoresist is coated, RIE (reactive ion etching) or wet etching technology is adopted to transfer a pattern to the mask material after lithography, or a mask layer material is deposited on the photoresist after the lithography step and then the photoresist is removed.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a schematic structural diagram of a fluorescent nanoscaler obtained in example 1 of the present invention;
FIG. 2 is a flow chart of a process for preparing the fluorescent nanoscaler in example 1 of the present invention;
FIG. 3 is a flow chart of a process for preparing the fluorescent nanoscaler in example 4 of the present invention;
FIG. 4 is a flow chart of a process for preparing the fluorescent nanoscaler in example 7 of the present invention;
FIG. 5 is a flow chart of a process for preparing the fluorescent nanoscaler in example 8 of the present invention;
FIG. 6 is a scanning electron microscope photograph of the nano-wire structure of the fluorescent nanoscaler component according to example 1 of Experimental example 1 of the present invention;
FIG. 7 is a fluorescence image of a fluorescent nanoscaler part according to example 4 of Experimental example 2 of the present invention;
FIG. 8 is a fluorescence image of a fluorescent nanoscaler part according to example 6 of Experimental example 3 of the present invention.
Description of reference numerals:
1-a substrate; 2-a mask layer; 21-a groove; 31-HSQ glue film; 32-conductive layer (espace 300Z); 33-mask layer (Cr); 41-mask layer (Ti); 42-ZEP 520A glue film; a 51-Si film; 52-cover glass; 53-ZEP 520A glue film layer; 54-Al conductive layer; 61-Au (gold); 62-Si film.
Detailed Description
The following examples are provided to further understand the present invention, not to limit the scope of the present invention, but to provide the best mode, not to limit the content and the protection scope of the present invention, and any product similar or similar to the present invention, which is obtained by combining the present invention with other prior art features, falls within the protection scope of the present invention.
The examples do not show the specific experimental steps or conditions, and can be performed according to the conventional experimental steps described in the literature in the field. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.
TABLE 1 reagents and apparatus used in the examples
| Reagents and materials | Source and model |
| Acetone (II) | Chinese medicine and analytical purity |
| Ethanol | Chinese medicine and analytical purity |
| HSQ glue | Dow kang Ning, XR1541-006 |
| ZEP520 | Japanese Raynaud, ZEP520A |
| Conductive polymer Escaper | Showa Denko, Japan, 300Z |
| TMAH | Chinese medicine and analytical purity |
| NMP | Chinese medicine and analytical purity |
| IPA | Chinese medicine and analytical purity |
| Reactive plasma etching machine | Oxford Plasmalab 80Plus |
| Electron beam exposure stage | Japanese JEOL JBX-5500ZA |
Example 1
The embodiment provides a method for preparing a fluorescent nanometer scale component, the process flow is shown in fig. 2, and the method comprises the following steps:
treating the substrate: sequentially subjecting a cover glass with the thickness of 0.17mm to ultrasonic treatment (40KHz) for 5 minutes by using acetone, ethanol and deionized water, and then treating for 5 minutes by using oxygen plasma under the power of 100W;
coating a photoresist: coating the HSQ glue with the mass fraction of 6% on a cover glass at 2000rpm for 40 seconds to obtain an HSQ glue film 31 with the thickness of 200nm, and baking the HSQ glue film 31 on a hot plate at 80 ℃ for 30 seconds to obtain an HSQ glue layer with moderate adhesive force; conducting polymer Espacener 300Z is spin-coated for 30 seconds at the rotating speed of 3000rpm, and then is placed on a hot plate at the temperature of 80 ℃ to be baked for 1 minute, so that a conducting layer (Espacener 300Z)32 is obtained;
photoetching: performing photoetching by adopting an electron beam with the voltage of 100kV, developing for 1min by using a mixed solution containing 1% by mass of NaOH and 4% by mass of NaCl, then washing for 30 seconds by using deionized water and IPA, and blow-drying by using nitrogen gas to form photoetching patterns of HSQ (high-speed polyethylene) glue nano line structures arranged at intervals on the substrate, wherein the transverse section width of each photoresist nano line structure is 10 nm;
transferring the pattern: depositing Cr as a mask layer (Cr)33 with a thickness of 100nm on the side of the substrate having the photolithographic pattern at a speed of 0.2 nm/s; and (3) immersing the sample into an HF aqueous solution with the mass fraction of 1% for treatment for 20 seconds, and corroding and removing the HSQ adhesive nano line structure to obtain the fluorescent nano scale component.
The structural schematic diagram of the fluorescence nanometer scale component is shown in fig. 1, and the fluorescence nanometer scale component comprises a substrate 1 and a mask layer 2 formed on the substrate, wherein the thickness of the mask layer 2 is 100nm, the mask layer 2 comprises grooves 21 arranged at intervals, the mask size of the grooves 21 is 10nm, and the depth-to-width ratio is 10.
Example 2
This example provides a specific implementation of a fluorescence nanoscale component, including the following steps:
treating the substrate: sequentially subjecting a cover glass with the thickness of 0.17mm to ultrasonic treatment (40KHz) for 10 minutes by using acetone, ethanol and deionized water, and then treating for 2 minutes by using oxygen plasma under the power of 50W;
coating a photoresist: coating an electron beam photoresist HSQ with the concentration of 1% on a substrate at the rotating speed of 4000rpm to obtain an HSQ adhesive film with the thickness of 100nm, and then placing the HSQ adhesive film on a hot plate with the temperature of 150 ℃ for baking for 20 seconds to obtain an HSQ adhesive layer with moderate adhesive force; depositing a layer of 20nm metal Al as a conductive layer;
photoetching: performing photoetching by adopting an electron beam with the voltage of 50kV, developing for 1 minute by using TMAH with the concentration of 25%, then flushing for 30 seconds by using deionized water and IPA, and then drying by using nitrogen;
transferring the pattern: depositing metal Ta with the thickness of 20nm at the speed of 0.1nm/s as a mask layer; and immersing the sample into HF aqueous solution with the concentration of 0.1% for treatment for 10 seconds, and corroding and removing the HSQ adhesive layer structure to obtain the fluorescent nano scale component, wherein the thickness of the mask layer is 20nm, the cross section width of the groove is 50nm, and the depth-to-width ratio is 0.4.
Example 3
This example provides a specific implementation of a fluorescence nanoscale component, including the following steps:
treating the substrate: sequentially subjecting a cover glass with the thickness of 0.17mm to ultrasonic treatment (40KHz) for 8 minutes by using acetone, ethanol and deionized water, and then treating for 5 minutes by using 80W oxygen plasma;
coating a photoresist: coating an electron beam photoresist HSQ with the concentration of 12% on a substrate at the rotating speed of 5000rpm to obtain an HSQ adhesive film with the thickness of 20nm, and then placing the HSQ adhesive film on a hot plate with the temperature of 50 ℃ for baking for 5 minutes; obtaining an HSQ adhesive layer with moderate adhesive force; depositing a layer of 20nm metal Cr as a conductive layer;
photoetching: carrying out photoetching by adopting an electron beam with the voltage of 30kV, and developing a sample for 1 minute by adopting TMAH with the concentration of 25%;
transferring the pattern: depositing metal Mo with the thickness of 50nm at the speed of 0.5nm/s as a mask layer; and immersing the sample into HF aqueous solution with the concentration of 0.2% for treatment for 60 seconds, and corroding and removing the HSQ adhesive layer structure to obtain the fluorescent nano scale component, wherein the thickness of the mask layer is 50nm, the cross section width of the groove is 150nm, and the depth-to-width ratio is 0.33.
Example 4
This example provides a specific implementation of a fluorescence nanoscale component, as shown in fig. 3, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm by using acetone, ethanol and deionized water for 8 minutes respectively, and then treating the cover glass by using 80W oxygen plasma for 3 minutes;
mask layer: depositing metallic Ti as a mask layer (Ti)41 with a thickness of 20nm on the substrate at a speed of 0.3 nm/s;
coating a photoresist: coating an electron beam photoresist ZEP520A 42 on the mask layer at the rotating speed of 4000rpm to obtain a ZEP520A glue film 42 with the thickness of 200nm, and then placing the film on a hot plate with the temperature of 120 ℃ for baking for 5 minutes;
photoetching: photoetching by adopting an electron beam with the voltage of 100kV, developing for 1 minute by using a developing solution ZED-N50 special for ZEP520A, and then blow-drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of each groove is 200 nm;
transferring the pattern: transferring the photoetching pattern to a mask layer (Ti) by using a reactive ion etching method (RIE, chlorine, carbon tetrafluoride) to form a metal nano-groove structure; and removing the residual glue of the ZEP by utilizing NMP or oxygen plasma to obtain the fluorescent nano scale component, wherein the thickness of the mask layer is 20nm, the cross section width of the groove is 200nm, and the depth-to-width ratio is 0.1.
Example 5
This example provides a specific implementation of a fluorescence nanoscale component, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm by using acetone, ethanol and deionized water for 8 minutes respectively, and then treating the cover glass by using 100W oxygen plasma for 3 minutes;
mask layer: depositing metallic Pd as a mask layer (Pd) with a thickness of 100nm on the substrate at a speed of 0.5 nm/s;
coating a photoresist: coating an electron beam photoresist ZEP520A on the mask layer at the rotating speed of 6000rpm to obtain a photoresist film layer (ZEP 520A) with the thickness of 300nm, and then placing the photoresist film layer on a hot plate at the temperature of 120 ℃ for baking for 5 minutes;
photoetching: carrying out photoetching by adopting an electron beam with the voltage of 100kV, developing for 1 minute by using a developing solution ZED-N50 special for ZEP520A, and then blowing and drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of each groove is 20 nm;
transferring the pattern: transferring the electron beam photoresist pattern to a mask layer (Pd) by RIE to form a metal nano-trench structure; and removing the residual glue of the ZEP by utilizing NMP or oxygen plasma to obtain the fluorescent nano scale component, wherein the thickness of the mask layer is 100nm, the cross section width of the groove is 20nm, and the depth-to-width ratio is 5.
Example 6
This example provides a specific implementation of a fluorescence nanoscale component, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm by using acetone, ethanol and deionized water for 8 minutes respectively, and then treating the cover glass by using 50W oxygen plasma for 3 minutes;
mask layer: depositing metallic Cr as a mask layer (Cr) on the substrate at a speed of 0.2nm/s to a thickness of 80 nm;
coating a photoresist: coating an electron beam photoresist ZEP520A on the mask layer at the rotating speed of 6000rpm to obtain a photoresist film layer (ZEP 520A) with the thickness of 500nm, and then placing the photoresist film layer on a hot plate at the temperature of 120 ℃ for baking for 5 minutes;
photoetching: carrying out photoetching by adopting an electron beam with the voltage of 100kV, developing for 1 minute by using a developing solution ZED-N50 special for ZEP520A, and then drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist film layer, wherein the cross section width of each groove is 20 nm;
transferring the pattern: transferring the electron beam photoresist pattern to the metal layer by RIE to form a metal nano-groove structure; and removing the residual glue of the ZEP by utilizing NMP or oxygen plasma to obtain the fluorescent nano scale component, wherein the thickness of the mask layer is 80nm, the width of the cross section of the groove is 20nm, and the depth-to-width ratio is 4.
Example 7
This example provides a specific implementation of a fluorescence nanoscale component, as shown in fig. 4, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm for 10 minutes by using acetone, ethanol and deionized water respectively, and then treating for 5 minutes by using 100W oxygen plasma for later use;
mask layer: an SOI substrate with a top Si film 51 with the thickness of 100nm is adopted, acetone, ethanol and deionized water are sequentially used for ultrasonic treatment for 5 minutes respectively, and then 100W oxygen plasma is used for treatment for 2 minutes; immersing the SOI wafer into a 20% HF aqueous solution for treatment for 16 hours, then fully cleaning the SOI wafer with deionized water, and inserting a treated cover glass 52 into the bottom of an Si film 51, wherein the Si film 51 is used as a mask layer; taking out the Si film 51 from the water by using a cover glass 52, drying and curing;
coating a photoresist: coating the electron beam photoresist ZEP520A on the mask layer at the rotating speed of 4000rpm to obtain a ZEP520A film layer 53 with the thickness of 250nm, and then placing the film layer on a hot plate with the temperature of 120 ℃ for baking for 5 minutes; depositing a layer of 20nm metal Al on the ZEP520A adhesive film layer 53 to serve as an Al conductive layer 54;
photoetching: carrying out photoetching by adopting an electron beam with the voltage of 100kV, developing for 1 minute by using a developing solution ZED-N50 special for ZEP520A, and then drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of the grooves is 150 nm;
pattern transfer: transferring the electron beam photoresist pattern to a silicon film by RIE (reactive ion etching) to form a Si film-based nano-groove structure; and removing residual glue of the ZEP520A by utilizing NMP or oxygen plasma to obtain the fluorescent nanometer scale component, wherein the thickness of the Si mask layer is 100nm, the cross section width of the groove is 150nm, and the depth-to-width ratio is 0.66.
Example 8
This example provides a specific implementation of a fluorescence nanoscale component, as shown in fig. 5, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm for 10 minutes by using acetone, ethanol and deionized water respectively, and then treating for 5 minutes by using 100W oxygen plasma for later use;
mask layer: an SOI substrate with the top layer Si thickness of 200nm is adopted, acetone, ethanol and deionized water are sequentially used for ultrasonic treatment for 10 minutes respectively, and then 50W oxygen plasma is used for treatment for 5 minutes; immersing the SOI wafer into a 20% HF aqueous solution for treatment for 16 hours, then fully cleaning the SOI wafer by using deionized water, and then inserting a cover glass into the bottom of the Si film 62, wherein the Si film 62 is used as a mask layer; fishing out the Si film from the water by using a cover glass, and drying and curing;
coating a photoresist: coating an electron beam photoresist ZEP520A on the mask layer at the rotating speed of 6000rpm to obtain a ZEP520A film layer with the thickness of 300nm, and then placing the film layer on a hot plate with the temperature of 120 ℃ for baking for 5 minutes; conducting polymer Escaper is coated on the photoresist film layer for 30 seconds in a spinning mode at the rotating speed of 3000rpm, and then the photoresist film layer is placed on a hot plate at the temperature of 80 ℃ to be baked for 1 minute;
photoetching: carrying out photoetching by adopting an electron beam with the voltage of 30kV, developing for 1 minute by using a special developing solution of ZEP520A, and then drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the width of the cross section of each groove is 10 nm;
pattern transfer: au (gold) 61 with the thickness of 40nm is deposited at the speed of 0.5nm/s, and then the Au (gold) is soaked in NMP at the temperature of 60 ℃ for 6 hours for stripping (lift-off) and glue removal; the re-immersion included HF at a concentration of 2M and H at a concentration of 0.1M2O2The mixed solution is treated for 30 minutes to carry out metal auxiliary corrosion, and finally the substrate is immersed into a gold corrosive solution to be treated for 10 minutes to remove a metal gold structure, so that the fluorescent nanometer scale component is obtained, wherein the thickness of the Si film mask layer is 200nm, the cross section width of the groove is 10m, and the depth-to-width ratio is 20.
Example 9
This example provides a specific implementation of a fluorescence nanoscale component, including the following steps:
treating the substrate: sequentially performing ultrasonic treatment on a cover glass with the thickness of 0.17mm by using acetone, ethanol and deionized water for 8 minutes respectively, and then treating the cover glass by using 80W oxygen plasma for 3 minutes;
mask layer: 2g of graphite powder was dispersed in 10g of a polyvinyl alcohol solution, spin-coated on a substrate at 3000rpm, and then baked on a hot plate at 120 ℃ for 5 minutes to obtain a carbon-doped polymer film having a thickness of 150nm on the substrate.
Coating a photoresist: coating electron beam photoresist ZEP520A on a carbon-doped polymer film at the rotating speed of 3000rpm to obtain a 300 nm-thick photoresist film layer, and then baking the photoresist film layer on a hot plate at the temperature of 120 ℃ for 5 minutes; conducting polymer Escaper 300Z is coated on the photoresist film layer for 30 seconds in a spinning mode at the rotating speed of 3000rpm, and then the photoresist film layer is placed on a hot plate at the temperature of 80 ℃ to be baked for 1 minute, so that the conducting layer is obtained.
Photoetching: carrying out photoetching by adopting an electron beam with the voltage of 100kV, developing for 1 minute by using a developing solution ZED-N50 special for ZEP520A, and then drying by using nitrogen gas to form photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of each groove is 100 nm;
transferring the pattern: transferring the electron beam photoresist pattern to a carbon-doped polymer film by using a reactive ion etching method (RIE, oxygen) to form a carbon-doped polymer nano groove structure, and obtaining the fluorescent nano scale component, wherein the thickness of the mask layer is 150nm, the cross section width of the groove is 100nm, and the depth-to-width ratio is 1.5.
Example 10
This example provides a specific embodiment of a fluorescent nanoscopic member, substantially identical to that described in example 8, except that gold is deposited at a thickness of 10nm at a rate of 0.1nm/s, and immersion is carried out in a bath comprising HF at a concentration of 2M and H at a concentration of 0.1M2O2The mixed solution is treated for 10 minutes to carry out metal auxiliary corrosion, and finally, the substrate is immersed in the gold corrosive solution to be treated for 1 minute to obtain the metal-based composite material.
Example 11
This example provides a specific embodiment of a fluorescent nanoscopic member, substantially identical to that described in example 8, except that gold is deposited at a thickness of 20nm at a rate of 0.2nm/s, and immersion is carried out in a bath comprising HF at a concentration of 2M and H at a concentration of 0.1M2O2The mixed solution is treated for 15 minutes to carry out metal auxiliary corrosion, and finally, the substrate is immersed in the gold corrosive solution to be treated for 3 minutes to obtain the metal-based composite material.
Comparative example 1
This comparative example provides a specific embodiment of a fluorescent nanoscaler component comprising the steps of:
the method comprises the following steps of (1) cleaning the surface of a glass sheet by using a transparent cover glass with the thickness of 18mm multiplied by 18mm as a substrate and sequentially using a concentrated chromic acid washing solution and a plasma cleaning machine; plating a layer of metal aluminum film with the thickness of 100nm on the surface of the cleaned glass sheet in a magnetron sputtering mode; plating a layer of Si with the thickness of 60nm on the non-transparent conducting layer3N4A mask layer; spin-coating a 90nm thick ZEP520(ZEON, Tokyo, JAPAN) photoresist on the surface of the mask layer at 4000rpm, then mounting the photoresist on an electron beam exposure platform (Vistec EBPG5000+ ES, Jane, Germany), exposing according to a preset pattern and developing with n-amyl acetate; etching with different gases twice through a plasma etcher (ICP-RIE SI500, Sentech, Berlin, Germany), wherein the first etching is performed by (SF)6,CHF3,O2) Mixed gas etching of Si3N4A mask layer, a second etching with (Cl)2,BCl3And N2) And etching the conductive aluminum film layer by using the mixed gas to obtain the fluorescent nanometer scale component with the mask layer thickness of 60nm and the groove section width of 200 nm.
Experimental example 1
In this experimental example, the fluorescent nanoscaler part obtained in example 1 was scanned by a scanning electron microscope (Hitachi S4800) to obtain an image as shown in fig. 6.
Experimental example 2
In this example, a fluorescence dye molecule Alexa Fluor 488 was used, 0.5mg of the fluorescein was dissolved in 5mL of PMMA (5%, ethyl acetate), coated on the fluorescent nanoscale member described in example 4 at 3000rpm, and an image was obtained by excitation at 488nm using a lecai fluorescence confocal microscope (Leica TCS SP8), as shown in fig. 7.
Experimental example 3
Using the fluorescent dye molecule Alexa Fluor 647, 0.5mg of this fluorescein was dissolved in 5mL of PMMA (5%, ethyl acetate), coated onto the fluorescent nanoscaler parts described in example 6 at 3000rpm, and the image obtained using a Leica fluorescence confocal microscope (Leica TCS SP8) under excitation at 647nm was shown in FIG. 8.
The description is given for clarity of illustration only and is not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (8)
1. A preparation method of a fluorescent nanometer scale component is characterized in that the fluorescent nanometer scale component comprises a substrate (1) and a mask layer (2) formed on the substrate, the mask layer comprises grooves (21) arranged at intervals, the cross section width of the grooves is 10-200nm, and the mask layer (2) is made of Si;
the preparation method comprises the following steps:
treating the substrate: cleaning the surface of the substrate;
mask layer: etching an SOI substrate by HF (hydrogen fluoride), carrying out water phase transfer on a Si film, and attaching Si on the substrate to be used as a mask layer material;
coating a photoresist: coating photoresist on the mask layer to form a photoresist mold layer with a thickness of 20-500 nm;
photoetching: patterning the photoresist mold layer, and forming photoetching patterns of grooves arranged at intervals on the photoresist mold layer, wherein the cross section width of each groove is 10-200 nm;
transferring the pattern: transferring the photoetching pattern of the photoresist film layer to the mask layer, and then removing the photoresist;
before the photoetching step, coating a layer of deposited conductive metal layer on the photoresist; the material for depositing the conductive metal layer is metal Al.
2. A method of manufacturing a fluorescent nanoscaler component according to claim 1, wherein the grooves are arranged at equal intervals, the cross-sectional width of the grooves being 10 to 150 nm; the depth-to-width ratio of the groove is 0.1-20.
3. A method of fabricating a fluorescent nanoscaler component according to claim 1, wherein the mask layer has a thickness of 100-200 nm.
4. A method of manufacturing a fluorescent nanoscaler component according to any one of claims 1 to 3, wherein the photoresist layer is patterned by electron beam lithography at a voltage of 30kV to 100 kV.
5. A method of manufacturing a fluorescent nanoscaler component according to any one of claims 1 to 3, wherein in the pattern transfer step, a photoresist pattern is transferred onto the mask layer using RIE or wet etching techniques.
6. A method of making a fluorescent nanoscaler component according to claim 5, wherein the wet etching step comprises a gold assisted etching step.
7. A fluorescent nanoscaler member produced by the production method according to any one of claims 1 to 6.
8. Use of the preparation method of any one of claims 1 to 6 or the fluorescent nanoscaled part prepared by the preparation method of any one of claims 1 to 6 in the field of microscopic fluorescence analysis.
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