CN110095441A - A kind of fluorescence nano scale member and its preparation and application - Google Patents

Abstract

The present invention relates to the technical field of ultra-high resolution fluorescent microscopic imaging, in particular to a fluorescent nanoscale component and its preparation and application; the fluorescent nanoscale component of the present invention includes a base layer and a mask layer, and the mask layer Grooves are arranged at intervals on the corresponding positions, the cross-sectional width of the grooves is 10-200nm, and the size (width) of the mask grooves is small, so as to ensure that the fluorescent nanoscales assembled by the fluorescent nanoscale components are accurate and can meet the requirements for ultra- The resolution of a high-resolution fluorescence microscope is required for calibration measurements.

Description

A kind of fluorescent nano-ruler component and its preparation and application

technical field

The invention relates to the technical field of ultra-high resolution fluorescent microscopic imaging, in particular to a fluorescent nano scale component and its preparation and application.

Background technique

Fluorescence microscopy plays an important role in the research of life sciences. It can study the life process and mechanism in the cell at the subcellular level. However, due to the limitation of the Abbe diffraction limit in traditional optics, its imaging resolution cannot exceed 200nm. the following. With the rise of ultra-high resolution fluorescence microscopy techniques, such as photosensitive localization microscopy (PALM), fluorescence photosensitive localization microscopy (FPALM), stochastic optical reconstruction microscopy (STORM), stimulated emission depletion microscopy, etc. (STED), Structured Illumination Microscopy (SIM) and Single Molecule Positioning Microscopy, etc. These techniques enable fluorescence microscopy to observe extremely fine structures from 20nm to 100nm.

With the continuous advancement and commercialization of ultra-high-resolution fluorescence microscopy technology, more and more universities, research institutes and companies have begun 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, it is often necessary to calibrate the resolution of the fluorescence microscope, so it is necessary to use a fluorescent standard structure with a defined size. In this regard, the Chinese patent document CN103712965A firstly proposed an equidistant fluorescent nano standard plate based on nano-grooves filled with fluorescent quantum dots. Although its equidistant structural form is more in line with the requirements of resolution measurement, its processing method is relatively cumbersome. , such as the first method described therein has too many preparation steps, especially the prepared nanowire structure cannot be formed at one time, and the quantum dot nanowire structure needs to be assembled multiple times to obtain a quantum dot nanowire structure with a certain aspect ratio. The steps are as follows: the first step is to spin-coat the photoresist layer 104 on the substrate 100, the second step is electron beam lithography to form an equidistant photoresist line structure, so that the substrate 100 is exposed, and the third step is to coat the exposed substrate 100 The charged polyelectrolyte polymer is adsorbed to form 109 layers. The fourth step is to use the Laber-by-layer assembly technology to form a multi-layer quantum dot nanowire structure 107 on the surface of the 109 layer. The quantum dot nanowire structure 107 needs to be between 3-10 layers , the fifth step is to remove the photoresist line structure, and the sixth step is to spin-coat a polymer layer or other protective layer 108 on the surface of layer 107 .

In order to solve the problem of cumbersome preparation steps of equidistant fluorescent nano standard plates, Chinese patent document CN103954600A proposes a fluorescent nanoscale structure with wide lines and narrow intervals based on filling fluorescent dyes in the trenches, but the structural form of wide lines and narrow intervals It does not meet the measurement standard of resolution and can only be used for general debugging and observation of fluorescence microscopes, and it uses photoresist as a mask, because ordinary photoresist cannot achieve such a high aspect ratio as 30nm wide and 100nm thick , leading to the limitation of the size of the prepared trench. In order to solve the above problems, in the preparation scheme, the non-transparent conductive layer is firstly reserved on the substrate, and then a layer of Si ₃ N ₄ with strong etching resistance is plated on it. , then spin-coat the photoresist, and finally expose the photoresist according to the predetermined pattern to dissolve the photoresist and sequentially etch the Si ₃ N ₄ layer and the non-transparent conductive layer. Compared with the groove structure with a large depth-width ratio, the scale structure size is large, the scale is not accurate, the mask layer thickness is small, and the light-shielding effect is poor, and because the non-transparent conductive layer under the mask layer has a low melting point such as an aluminum film, It is unstable under high-energy laser conditions, prone to deformation, and has a short service life.

Contents of the invention

Therefore, the technical problem to be solved by the present invention is to overcome the inaccurate defect of the fluorescent nano-scale parts in the prior art, and then provide a kind of high-precision fluorescent nano-scale parts and the method and method for preparing the fluorescent nano-scale parts disclosed in the present invention. application.

To this end, the present invention provides a fluorescent nanoscale component, which includes a substrate and a mask layer formed on the substrate, the mask layer includes grooves arranged at intervals, and the cross-sectional width of the grooves is 10-200 nm.

Preferably, the grooves are arranged at equal intervals, and the cross-sectional width of the grooves is 10-150 nm; preferably, the aspect ratio of the grooves is 0.1-20.

Preferably, the material of the mask layer is selected from high melting point metals or non-metallic materials that cannot transmit ultraviolet and visible light.

Preferably, the metal with a high melting point is selected from one of Ta, Mo, Cr, Ti and Pd; the non-metallic material that cannot transmit ultraviolet and visible light is Si or a high-density carbon-doped polymer.

Preferably, when the material of the mask layer is a high melting point metal, the thickness of the mask layer is 20-100nm; when the material of the mask layer is Si, the thickness of the mask layer is 100-200nm .

The present invention also discloses a preparation method of the fluorescent nano scale part, comprising the following steps:

Treating the substrate: cleaning the surface of the substrate;

Mask layer: formed on the surface of the substrate;

Coating photoresist: coating the photoresist on the mask layer to form a photoresist mold layer with a thickness of 20-500nm;

Photolithography: patterning the photoresist mold layer, forming photolithographic patterns of grooves arranged at intervals on the photoresist mold layer, and the cross-sectional width of the grooves is 10-200nm;

Transferring the pattern: transferring the photoresist pattern of the photoresist mold layer to the mask layer, and then removing the photoresist.

The present invention also discloses a preparation method of the fluorescent nano scale part, comprising the following steps:

Treating the substrate: cleaning the surface of the substrate;

Coating photoresist: coating the photoresist on the substrate to form a photoresist mold layer with a thickness of 20-500nm;

Photolithography: patterning the photoresist mold layer, forming a photoresist pattern of photoresist nanowire structures arranged at intervals on the substrate, the cross-sectional width of the photoresist nanowire structure is 10 -200nm;

Transferring the pattern: depositing a mask layer material on the side of the substrate having the photoresist pattern to form a mask layer, then removing the photoresist, and transferring the photoresist pattern to the mask layer.

Preferably, the photoresist mold layer is patterned by electron beam lithography, and the voltage of the electron beam lithography is 30kV-100kV.

Preferably, when the high melting point metal is used as the material of the mask layer, it is deposited on the substrate by deposition, sputtering or electron beam evaporation, and the deposition rate is 0.1-0.5 nm/s.

Preferably, when Si is used as the material of the mask layer, before coating the photoresist, the SOI substrate is etched with HF, and the Si film is transferred in water, and Si is attached to the substrate as the material of the mask layer.

Preferably, in the step of transferring the pattern, the photoresist pattern is transferred to the mask layer by RIE or wet etching technology.

Preferably, the wet etching step includes a gold assisted etching step.

Preferably, before the photolithography step, a conductive polymer layer or a conductive metal layer is deposited on the photoresist.

The invention also discloses an application of the fluorescent nanoscale part or the fluorescent nanoscale part prepared by the method for preparing the fluorescent nanoscale part in the field of microscopic fluorescence analysis.

The technical solution of the present invention has the following advantages:

1. The fluorescent nanoscale ruler part of the present invention comprises a substrate and a mask layer formed on the substrate, grooves are arranged at intervals on the corresponding positions of the mask layer, and the cross-sectional width of the groove is 10-200nm, and the mask groove The size (width) of the groove is small, so as to ensure the accuracy of the fluorescent nanoscale assembled by the fluorescent nanoscale component, which can meet the requirements for calibration and measurement of the resolution of the ultra-high resolution fluorescence microscope.

2. The fluorescent nano-scale parts of the present invention, the grooves are arranged at equal intervals, and the cross-sectional width of the grooves is 10-150nm; the grooves are arranged at equal intervals, which overcomes the existing fluorescent nano-scale parts The structural form of medium-wide lines and narrow intervals leads to defects that do not meet the resolution measurement standard of the microscope, and can meet the requirements for calibration measurement of the resolution of the super-resolution fluorescence microscope; the cross-sectional width of the groove is 10-150nm, further Improve the accuracy of fluorescent nanoscale;

Further, the aspect ratio of the trench is 0.1-20, the thickness of the trench mask is further controlled to be 20-200nm, and the aspect ratio can reach up to 200nm/10nm at most, the light blocking effect is good, and the scale accuracy is high .

3. The material of the mask layer of the fluorescent nano scale part of the present invention is selected from high-melting point metals, Si, and high-density carbon-doped polymers. When in use, it has better stability and will not be affected by high-energy lasers. Deformation occurs due to long-term irradiation, thereby prolonging the service life; since high-melting point metals are conductors, Si and high-density carbon-doped polymers are semiconductors, and there is no need to use conductive material layers in the process, only the base and mask layers, the process Simple.

4. The preparation method of the fluorescent nano-scale part of the present invention adopts the mode of electron beam lithography, and before coating photoresist, high-melting point metal mask material or Si film are attached on the substrate, adopt RIE or wet after photolithography The pattern is transferred to the mask material by etching technology, or the mask layer material is deposited on the photoresist after the photolithography step and then the photoresist is removed. This preparation method can be flexibly adjusted according to the mask material. The obtained phosphor lines in the mask groove can be single lines, equidistant lines, wide lines and narrow intervals, so as to meet different usage requirements, and during the preparation process, it is possible to prevent the conductive layer from remaining on the fluorescent nanoscale parts and improve the mask The function of the structure.

Description of drawings

In order to more clearly illustrate the specific implementation of the present invention or the technical solutions in the prior art, the following will briefly introduce the accompanying drawings that need to be used in the specific implementation or description of the prior art. Obviously, the accompanying drawings in the following description The drawings show some implementations of the present invention, and those skilled in the art can obtain other drawings based on these drawings without any creative work.

Fig. 1 is the structural representation of fluorescent nano scale obtained in the embodiment 1 of the present invention;

Fig. 2 is a process flow diagram for preparing the fluorescent nanoscale in Example 1 of the present invention;

Fig. 3 is the process flow diagram of preparing described fluorescent nano scale in the embodiment of the present invention 4;

Fig. 4 is the process flow diagram of preparing described fluorescent nano scale in the embodiment of the present invention 7;

Figure 5 is a process flow diagram for preparing the fluorescent nanoscale in Example 8 of the present invention;

Fig. 6 is a scanning electron microscope picture of the nanowire structure of the fluorescent nanoscale member described in Example 1 in Experimental Example 1 of the present invention;

Fig. 7 is the fluorescent imaging diagram of the fluorescent nanoscale member described in Example 4 in Experimental Example 2 of the present invention;

Fig. 8 is a fluorescence imaging diagram of the fluorescent nanoscale member described in Example 6 in Experimental Example 3 of the present invention.

Explanation of reference signs:

1-substrate; 2-mask layer; 21-groove; 31-HSQ adhesive film; 32-conductive layer (Espacer 300Z); 33-mask layer (Cr); 41-mask layer (Ti); 42- ZEP 520A film; 51-Si film; 52-cover glass; 53-ZEP 520A film layer; 54-Al conductive layer; 61-Au (gold); 62-Si film.

Detailed ways

The following examples are provided in order to further understand the present invention better, are not limited to the best implementation mode, and do not limit the content and protection scope of the present invention, anyone under the inspiration of the present invention or use the present invention Any product identical or similar to the present invention obtained by combining features of other prior art falls within the protection scope of the present invention.

If no specific experimental steps or conditions are indicated in the examples, it can be carried out according to the operation or conditions of the conventional experimental steps described in the literature in this field. The reagents or instruments used were not indicated by the manufacturer, and they were all commercially available conventional products.

Reagents and instruments used in the embodiment of table 1

Reagents and Materials source and model acetone Chinese medicine, analytically pure Ethanol Chinese medicine, analytically pure HSQ glue US Dow Corning, XR1541-006 ZEP520 Japan Zeon, ZEP520A Conductive Polymer Espacer Japan Showa Denko, 300Z TMAH Chinese medicine, analytically pure NMP Chinese medicine, analytically pure IPA Chinese medicine, analytically pure Reactive Plasma Etcher Oxford Plasmalab 80Plus Electron Beam Exposure Station Japan JEOL JBX-5500ZA

Example 1

This embodiment provides a method for preparing a fluorescent nanoscale member, the process flow is shown in Figure 2, including the following steps:

Treat the substrate: Sonicate (40KHz) the cover glass with a thickness of 0.17mm for 5 minutes with acetone, ethanol, and deionized water in sequence, and then treat it with oxygen plasma at 100W for 5 minutes;

Coating photoresist: Apply HSQ glue with a mass fraction of 6% on the cover glass at 2000rpm for 40 seconds to obtain a 200nm thick HSQ film 31, and bake it on a hot plate at 80°C for 30 seconds to obtain adhesion Moderate HSQ adhesive layer; Spin-coat the conductive polymer Espacer 300Z at a speed of 3000rpm for 30 seconds, and then bake it on a hot plate at 80°C for 1 minute to obtain a conductive layer (Espacer 300Z) 32;

Photolithography: photolithography was carried out with an electron beam at a voltage of 100kV, developed with a mixed solution containing 1% NaOH and 4% NaCl for 1 min, then rinsed with deionized water and IPA for 30 seconds, and then flushed with nitrogen gas Blow dry to form photolithographic patterns of HSQ glue nanowire structures arranged at intervals on the substrate, the cross-sectional width of the photoresist nanowire structure is 10nm;

Transfer pattern: Depositing Cr with a thickness of 100 nm as a mask layer (Cr) 33 at a speed of 0.2 nm/s on the side with the photolithography pattern on the substrate; immersing the sample into 1% HF aqueous solution by mass fraction After 20 seconds of medium treatment, the HSQ glue nanowire structure was etched and removed to obtain a fluorescent nanoscale component.

The schematic structural view of the fluorescent nanometer scale part is shown in Figure 1, including a substrate 1 and a mask layer 2 formed on the substrate, the thickness of the mask layer 2 is 100nm, and the mask layer 2 includes intervals. A groove 21, the mask size of the groove 21 is 10 nm, and the aspect ratio is 10.

Example 2

The present embodiment provides a specific implementation of the fluorescent nano scale part, comprising the following steps:

Treat the substrate: Sonicate (40KHz) the cover glass with a thickness of 0.17mm for 10 minutes with acetone, ethanol, and deionized water in sequence, and then treat it with oxygen plasma at 50W for 2 minutes;

Coating photoresist: apply electron beam photoresist HSQ with a concentration of 1% on the substrate at a speed of 4000rpm to obtain a HSQ film with a thickness of 100nm, and then bake it on a hot plate at a temperature of 150°C 20 seconds to get a HSQ adhesive layer with moderate adhesion; deposit a layer of 20nm metal Al as a conductive layer;

Photolithography: use an electron beam with a voltage of 50kV for photolithography, develop with TMAH with a concentration of 25% for 1 minute, then rinse with deionized water and IPA for 30 seconds, and then blow dry with nitrogen;

Transfer pattern: Deposit metal Ta with a thickness of 20nm at a rate of 0.1nm/s as a mask layer; immerse the sample in a 0.1% HF aqueous solution for 10 seconds, and remove the HSQ adhesive layer structure by etching to obtain a fluorescent nanoscale The thickness of the mask layer of the component is 20nm, the section width of the groove is 50nm, and the aspect ratio is 0.4.

Example 3

The present embodiment provides a specific implementation of the fluorescent nano scale part, comprising the following steps:

Treat the substrate: Sonicate (40KHz) each of acetone, ethanol, and deionized water for 8 minutes on the cover glass with a thickness of 0.17mm, and then treat it with 80W oxygen plasma for 5 minutes;

Coating photoresist: apply electron beam photoresist HSQ with a concentration of 12% on the substrate at a speed of 5000rpm to obtain a HSQ film with a thickness of 20nm, and then bake it on a hot plate at a temperature of 50°C 5 minutes; obtain a HSQ adhesive layer with moderate adhesion; deposit a layer of 20nm metal Cr as a conductive layer;

Photolithography: use an electron beam with a voltage of 30kV for photolithography, and use TMAH with a concentration of 25% to develop the sample for 1 minute;

Transfer pattern: Deposit metal Mo with a thickness of 50nm at a rate of 0.5nm/s as a mask layer; immerse the sample in a 0.2% HF aqueous solution for 60 seconds, and remove the HSQ glue layer structure by etching to obtain a fluorescent nanoscale For the component, the thickness of the mask layer is 50nm, the section width of the trench is 150nm, and the aspect ratio is 0.33.

Example 4

The present embodiment provides a specific embodiment of a fluorescent nanometer scale part, as shown in Figure 3, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm for 8 minutes with acetone, ethanol, and deionized water in sequence, and then treat it with 80W oxygen plasma for 3 minutes;

Mask layer: metal Ti with a thickness of 20 nm is deposited on the substrate at a rate of 0.3 nm/s as a mask layer (Ti) 41;

Coating photoresist: Apply electron beam photoresist ZEP 520A 42 on the mask layer at a speed of 4000rpm to obtain a ZEP 520A film 42 with a thickness of 200nm, and then place it on a hot plate at a temperature of 120°C for baking Bake for 5 minutes;

Photolithography: use an electron beam with a voltage of 100kV to carry out photolithography, develop with ZED-N50, a special developer solution for ZEP 520A, for 1 minute, and then blow dry with nitrogen to form the photoresist pattern of grooves arranged at intervals on the photoresist mold layer. Engraving patterns, the cross-sectional width of the trench is 200nm;

Transfer pattern: use reactive ion etching (RIE, chlorine, carbon tetrafluoride) to transfer the photolithographic pattern to the mask layer (Ti) to form a metal nano-groove structure; use NMP or oxygen plasma to remove ZEP residue , to obtain a fluorescent nanoscale member, the thickness of the mask layer is 20nm, the cross-sectional width of the groove is 200nm, and the aspect ratio is 0.1.

Example 5

The present embodiment provides a specific implementation of the fluorescent nano scale part, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm for 8 minutes each in acetone, ethanol, and deionized water, and then treat it with 100W oxygen plasma for 3 minutes;

Mask layer: metal Pd with a thickness of 100 nm is deposited on the substrate at a rate of 0.5 nm/s as a mask layer (Pd);

Coating photoresist: apply electron beam photoresist ZEP 520A on the mask layer at a speed of 6000rpm to obtain a photoresist film layer (ZEP 520A) with a thickness of 300nm, and then put it in a hot water at a temperature of 120°C Bake on board for 5 minutes;

Photolithography: use an electron beam with a voltage of 100kV to carry out photolithography, develop with ZED-N50, a special developer solution for ZEP 520A, for 1 minute, and then blow dry with nitrogen to form the photoresist pattern of grooves arranged at intervals on the photoresist mold layer. Engraving patterns, the cross-sectional width of the trench is 20nm;

Transfer pattern: Use RIE to transfer the electron beam photoresist pattern to the mask layer (Pd) to form a metal nano-groove structure; use NMP or oxygen plasma to remove the ZEP residue to obtain a fluorescent nanoscale part, the mask The layer thickness is 100 nm, the groove cross-sectional width is 20 nm, and the aspect ratio is 5.

Example 6

The present embodiment provides a specific implementation of the fluorescent nano scale part, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm for 8 minutes each in acetone, ethanol, and deionized water, and then treat it with 50W oxygen plasma for 3 minutes;

Mask layer: metal Cr with a thickness of 80 nm is deposited on the substrate at a rate of 0.2 nm/s as a mask layer (Cr);

Coating photoresist: apply electron beam photoresist ZEP 520A on the mask layer at a speed of 6000rpm to obtain a photoresist film layer (ZEP 520A) with a thickness of 500nm, and then put it in a hot water at a temperature of 120°C Bake on board for 5 minutes;

Photolithography: use an electron beam with a voltage of 100kV to carry out photolithography, use ZEP 520A special developer ZED-N50 to develop for 1 minute, and then blow dry with nitrogen to form the photoresist of grooves arranged at intervals on the photoresist film layer. Engraving patterns, the cross-sectional width of the trench is 20nm;

Transfer pattern: use RIE to transfer the electron beam photoresist pattern to the metal layer to form a metal nano-groove structure; use NMP or oxygen plasma to remove the ZEP residual glue to obtain fluorescent nanoscale components, and the thickness of the mask layer is 80nm , the cross-sectional width of the trench is 20 nm, and the aspect ratio is 4.

Example 7

This embodiment provides a specific implementation of a fluorescent nanometer scale part, as shown in Figure 4, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm with acetone, ethanol, and deionized water for 10 minutes, and then treat it with 100W oxygen plasma for 5 minutes, and set aside;

Mask layer: adopt the SOI substrate with the top Si film 51 thickness of 100nm, use acetone, ethanol, and deionized water to sonicate for 5 minutes each, and then treat it with 100W oxygen plasma for 2 minutes; immerse the SOI sheet in 20% HF treated in an aqueous solution for 16 hours, then fully cleaned with deionized water, and the treated cover glass 52 was inserted into the bottom of the Si film 51, and the Si film 51 was used as a mask layer; the Si film 51 was pulled out from the water with the cover glass 52 , dry and solidify;

Coating photoresist: Apply electron beam photoresist ZEP 520A on the mask layer at a speed of 4000rpm to obtain a ZEP 520A film layer 53 with a thickness of 250nm, and then place it on a hot plate at a temperature of 120°C for baking Bake for 5 minutes; deposit a layer of 20nm metal Al on the ZEP520A film layer 53 as the Al conductive layer 54;

Photolithography: use an electron beam with a voltage of 100kV to carry out photolithography, develop with ZED-N50, a special developer solution for ZEP 520A, for 1 minute, and then blow dry with nitrogen to form the photoresist pattern of grooves arranged at intervals on the photoresist mold layer. Engraving patterns, the cross-sectional width of the trench is 150nm;

Pattern transfer: Use RIE to transfer the electron beam photoresist pattern to the silicon film to form a nano-groove structure based on the Si film; use NMP or oxygen plasma to remove the ZEP 520A residual glue to obtain a fluorescent nanoscale part. The Si mask The thickness of the mold layer is 100 nm, the section width of the trench is 150 nm, and the aspect ratio is 0.66.

Example 8

The present embodiment provides a specific implementation of the fluorescent nano-ruler component, as shown in Figure 5, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm with acetone, ethanol, and deionized water for 10 minutes, and then treat it with 100W oxygen plasma for 5 minutes, and set aside;

Mask layer: use an SOI substrate with a thickness of 200nm on the top layer of Si, ultrasonically use acetone, ethanol, and deionized water for 10 minutes each, and then treat it with 50W oxygen plasma for 5 minutes; immerse the SOI sheet in an aqueous solution of 20% HF Treat in medium for 16 hours, then fully wash with deionized water, then insert the cover glass into the bottom of the Si film 62, and the Si film 62 is used as a mask layer; use the cover glass to remove the Si film from the water, dry and solidify;

Coating photoresist: apply electron beam photoresist ZEP 520A on the mask layer at a speed of 6000rpm to obtain a ZEP 520A film layer with a thickness of 300nm, and then bake it on a hot plate at a temperature of 120°C for 5 Minutes; Spin-coat the conductive polymer Espacer on the photoresist film layer at a speed of 3000rpm for 30 seconds, and then bake it on a hot plate at 80°C for 1 minute;

Photolithography: use an electron beam with a voltage of 30kV to carry out photolithography, develop with ZEP 520A special developer for 1 minute, and then blow dry with nitrogen to form a photolithographic pattern of grooves arranged at intervals on the photoresist mold layer, The cross-sectional width of the trench is 10nm;

Pattern transfer: Deposit Au (gold) 61 with a thickness of 40nm at a speed of 0.5nm/s, and then immerse in NMP at 60°C for 6 hours for lift-off removal; then immerse in 2M HF and Concentration is 0.1M H ₂ O ₂ in the mixed solution processing 30 minutes to carry out metal-assisted etching, finally immerse the substrate in the gold corrosion solution and treat for 10 minutes to remove the metal gold structure, obtain the fluorescent nano scale part, the Si film mask The layer thickness is 200 nm, the cross-sectional width of the trench is 10 m, and the aspect ratio is 20.

Example 9

The present embodiment provides a specific implementation of the fluorescent nano scale part, comprising the following steps:

Treat the substrate: Sonicate the cover glass with a thickness of 0.17mm for 8 minutes with acetone, ethanol, and deionized water in sequence, and then treat it with 80W oxygen plasma for 3 minutes;

Mask layer: Disperse 2g of graphite powder in 10g of polyvinyl alcohol solution, spin-coat it on the substrate at a speed of 3000rpm, and bake it on a hot plate at 120°C for 5 minutes to obtain a thickness on the substrate 150nm carbon-doped polymer film.

Coating photoresist: On the carbon-doped polymer film, apply the electron beam photoresist ZEP 520A at a speed of 3000rpm to obtain a photoresist film layer with a thickness of 300nm, and then put it in a hot water at a temperature of 120°C The board was baked for 5 minutes; the conductive polymer Espacer 300Z was spin-coated on the photoresist film layer at a speed of 3000 rpm for 30 seconds, and then placed on a hot plate at 80° C. for 1 minute to obtain a conductive layer.

Photolithography: use an electron beam with a voltage of 100kV to carry out photolithography, develop with ZED-N50, a special developer solution for ZEP 520A, for 1 minute, and then blow dry with nitrogen to form the photoresist pattern of grooves arranged at intervals on the photoresist mold layer. Engraving patterns, the cross-sectional width of the trench is 100nm;

Transfer pattern: use reactive ion etching method (RIE, oxygen) to transfer the electron beam photoresist pattern to the carbon-doped polymer film, form the carbon-doped polymer nano-groove structure, and obtain the fluorescent nanoscale part, the mask The thickness of the mold layer is 150nm, the section width of the trench is 100nm, and the aspect ratio is 1.5.

Example 10

This embodiment provides a specific implementation of a fluorescent nanometer scale component, which is basically the same as the implementation described in Example 8, the difference is that gold with a thickness of 10 nm is deposited at a speed of 0.1 nm/s, and gold with a concentration of 2M is immersed in Treat in a mixed solution of HF and 0.1M H ₂ O ₂ for 10 minutes to carry out metal-assisted etching, and finally immerse the substrate in a gold etching solution for 1 minute to obtain the finished product.

Example 11

This embodiment provides a specific implementation of a fluorescent nanoscale member, which is basically the same as the implementation described in Example 8, the difference is that gold with a thickness of 20 nm is deposited at a speed of 0.2 nm/s, and the immersion includes gold with a concentration of 2M. Treat in a mixed solution of HF and 0.1M H ₂ O ₂ for 15 minutes to carry out metal-assisted etching, and finally immerse the substrate in a gold etching solution for 3 minutes to obtain the finished product.

Comparative example 1

This comparative example provides a kind of specific embodiment of fluorescent nano scale part, comprises the following steps:

A 18mm×18mm transparent cover glass is used as the substrate, and the surface of the glass sheet is cleaned with a concentrated chromic acid solution and a plasma cleaner in sequence; a layer of 100nm thick is coated on the surface of the cleaned glass sheet by magnetron sputtering A metal aluminum film; a 60nm thick Si ₃ N ₄ mask layer is plated on the non-transparent conductive layer; a 90nm thick ZEP520 (ZEON, Tokyo, JAPAN) photoresist, then installed on the electron beam exposure platform (Vistec EBPG5000+ES, Jane, Germany), exposed according to the preset pattern and developed with n-pentyl acetate; SI500, Sentech, Berlin, Germany) were etched twice with different gases, wherein the first etching used (SF ₆ , CHF ₃ , O ₂ ) mixed gas to etch the Si ₃ N ₄ mask layer, and the second etching The conductive aluminum film layer was etched with (Cl ₂ , BCl ₃ and N ₂ ) mixed gas to obtain a fluorescent nanoscale member with a mask layer thickness of 60 nm and a groove cross-sectional width of 200 nm.

Experimental example 1

In this experimental example, the fluorescent nanoscale component obtained in Example 1 was scanned by a scanning electron microscope (Hitachi S4800) to obtain an imaging image as shown in FIG. 6 .

Experimental example 2

This experimental example uses the fluorescent dye molecule Alexa Fluor 488, takes 0.5 mg of the fluorescein and dissolves it in 5 mL of PMMA (5%, ethyl acetate), and coats it on the fluorescent nanoscale member described in Example 4 at a speed of 3000 rpm. The image obtained by fluorescence confocal microscope (Leica TCS SP8) under 488nm excitation is shown in Figure 7.

Experimental example 3

Using fluorescent dye molecule Alexa Fluor 647, take 0.5 mg of the fluorescein and dissolve it in 5 mL of PMMA (5%, ethyl acetate), and apply it to the fluorescent nanoscale member described in Example 6 at a speed of 3000 rpm, and use Leica fluorescent confocal The image obtained by a microscope (Leica TCS SP8) under 647nm excitation is shown in Figure 8.

It is merely an example for clear description, and does not limit the embodiment. For those of ordinary skill in the art, other changes or changes in different forms can be made on the basis of the above description. It is not necessary and impossible to exhaustively list all the implementation manners here. And the obvious changes or changes derived therefrom are still within the scope of protection of the present invention.

Claims (14)

1. a kind of fluorescent nano scale part, comprise the mask layer (2) that forms on substrate (1) and substrate, described mask layer comprises the groove (21) that is arranged at intervals, and the section width of described groove is 10 -200nm. 2. The fluorescent nanoscale member according to claim 1, wherein the grooves are arranged at equal intervals, and the cross-sectional width of the grooves is 10-150nm; preferably, the aspect ratio of the grooves is 0.1-20. 3. The fluorescent nanoscale member according to claim 1 or 2, characterized in that, the material of the mask layer is selected from high melting point metals or non-metallic materials that cannot transmit ultraviolet and visible light. 4. fluorescent nano-scale ruler parts according to claim 3, is characterized in that, described refractory metal is selected from the one in Ta, Mo, Cr, Ti and Pd; The non-metallic material that described ultraviolet-visible light can not transmit is Si or high density carbon doped polymers. 5. The fluorescent nanoscale ruler part according to any one of claims 1-4, wherein when the material of the mask layer is a refractory metal, the thickness of the mask layer is 20-100nm; When the material of the mask layer is Si, the thickness of the mask layer is 100-200 nm. 6. A method for preparing fluorescent nanoscale parts as claimed in any one of claims 1-5, characterized in that, comprising the following steps: Treating the substrate: cleaning the surface of the substrate; Mask layer: formed on the surface of the substrate; Coating photoresist: coating the photoresist on the mask layer to form a photoresist mold layer with a thickness of 20-500nm; Photolithography: patterning the photoresist mold layer, forming photolithographic patterns of grooves arranged at intervals on the photoresist mold layer, and the cross-sectional width of the grooves is 10-200nm; Transferring the pattern: transferring the photoresist pattern of the photoresist film layer to the mask layer, and then removing the photoresist. 7. A method for preparing fluorescent nano scale parts as claimed in any one of claims 1-5, characterized in that, comprising the following steps: Treating the substrate: cleaning the surface of the substrate; Coating photoresist: coating the photoresist on the substrate to form a photoresist film layer with a thickness of 20-500nm; Photolithography: patterning the photoresist mold layer, forming a photoresist pattern of photoresist nanowire structures arranged at intervals on the substrate, the cross-sectional width of the photoresist nanowire structure is 10 -200nm; Transferring the pattern: depositing a mask layer material on the side of the substrate having the photoresist pattern to form a mask layer, then removing the photoresist, and transferring the photoresist pattern to the mask layer. 8. according to the preparation method of claim 6 or 7 described fluorescent nano scale parts, it is characterized in that, adopt the mode of electron beam lithography to carry out patterning to described photoresist mold layer, the voltage of described electron beam lithography 30kV-100kV. 9. the preparation method of fluorescent nanoscale ruler parts according to claim 6 is characterized in that, when using refractory metal as mask layer material, by deposition, sputtering or electron beam evaporation on the substrate, deposition rate is 0.1- 0.5nm/s. 10. the preparation method of fluorescent nano-scale member according to claim 6 is characterized in that, when using Si as mask layer material, before coating photoresist, SOI substrate is corroded by HF, Si film water phase transfer , attaching Si on the substrate as a mask layer material. 11. according to claim 6, the preparation method of the described fluorescent nano-scale member of any one of 8-10, it is characterized in that, in described transfer pattern step, adopt RIE or wet etching technology to transfer photoresist pattern onto the mask layer. 12 . The method for preparing fluorescent nanoscale components according to claim 11 , wherein the wet etching step comprises a gold-assisted etching step. 13 . 13. according to the preparation method of the described fluorescent nano-ruler part according to any one of claim 6-12, it is characterized in that, before the photolithography step, also coat one deck conductive polymer layer or on the photoresist A conductive metal layer is deposited. 14. The application of the fluorescent nanoscale part according to any one of claims 1-5 or the fluorescent nanoscale part prepared by the method according to any one of claims 6-13 in the field of microscopic fluorescence analysis.