Grid structure for cryoelectron microscope and preparation method thereof

Abstract

The invention discloses a grid structure for a cryoelectron microscope and a preparation method of the grid structure. The grid structure for the cryoelectron microscope comprises a support body, a support film and an interlayer, the supporting film is provided with at least one through accommodating channel; the interlayer is located between the supporting film and the supporting body, an imaging window is defined by the interlayer and the hollow part of the supporting body, and the containing channel is located within the range of the imaging window. On one hand, in the etching process of the supporting film, the sacrificial layer is located at the bottom of the supporting film and plays a role in supporting and protecting the supporting film, and the situation that the vacuum degree of the grid structure is reduced due to supporting film hole forming is avoided; and on the other hand, a sacrificial layer is not arranged in some embodiments of the invention, so that the problem of high resistance caused by the sacrificial layer is improved, and the conductivity of the sample is remarkably improved. Meanwhile, in the preparation process of the supporting body, the first supporting sheet is located at the top of the supporting film and also plays a role in supporting and protecting the supporting film.

Description

Technical Field

[0001] This application relates to the field of cryo-electron microscopy, and in particular to a grid structure for cryo-electron microscopy and its preparation method. Background Technology

[0002] In cryo-electron microscopy, a support mesh with a porous structure is often used. This porous structure allows the sample solution to remain within the pores and form a film at those pores. When the electron beam emitted by the electron microscope passes through the sample, it completes the imaging of the sample.

[0003] In related technologies, the support membrane of the carrier is isolated and perforated, which is easily affected by the perforations, causing a decrease in vacuum and resulting in poor adsorption of the support membrane, thus affecting the yield. On the other hand, the process of bonding the support membrane to the substrate and then perforating it can ensure the vacuum of the product, but it is prone to problems such as reduced membrane structure flatness and warping.

[0004] In cryo-electron microscopy, the commonly used film structure materials have poor conductivity, which makes it easy for charge to accumulate on the sample surface when irradiated by electron beam, thus causing sample drift and a decrease in imaging quality. Meanwhile, metal layers that help with heat capacity are difficult to process on the screen.

[0005] Meanwhile, to improve the support of the support membrane, support beams are often set on the surface of the support membrane. However, thicker support beams will increase the thickness of the part where the liquid film contacts the beam, reducing the imaging and observation effect of the carrier mesh.

[0006] In addition, the current mesh structure is relatively thick. Although it can improve the overall strength of the mesh structure, it is not conducive to obtaining a thinner and more uniform ice layer in the later biological sample preparation, resulting in poor imaging effect of the mesh. Summary of the Invention

[0007] This application aims to at least solve one of the technical problems existing in the prior art. To this end, this application proposes a method for preparing a mesh structure, and also proposes a mesh structure for cryo-electron microscopy.

[0008] According to a first aspect embodiment of the present application, the method for preparing a carrier screen includes:

[0009] Prepare a substrate and then prepare a support film on top of the substrate;

[0010] The support film is etched to form at least one accommodating channel;

[0011] A first support sheet is disposed on top of the support membrane;

[0012] A support is prepared at the bottom of the support membrane.

[0013] The grid structure for cryo-electron microscopy according to the embodiments of this application has at least the following beneficial effects: On the one hand, during the etching process of the support film, the sacrificial layer is located at the bottom of the support film and plays a supporting and protective role for the support film, avoiding the decrease in vacuum degree of the grid structure caused by the pore making of the support film, and also avoiding damage and contamination of the film structure, ensuring the adsorption effect and flatness of the support film, and improving the yield; on the other hand, in some embodiments of this application, the sacrificial layer is not set, reducing the overall resistivity of the grid, improving the high resistance problem caused by the sacrificial layer, and significantly improving the conductivity of the sample. At the same time, during the preparation of the support, the first support sheet is located at the top of the support film, which also plays a supporting and protective role for the support film, avoiding the support film being affected by the preparation of the support.

[0014] According to some embodiments of this application, the preparation of the substrate and the preparation of a support film on top of the substrate includes:

[0015] A sacrificial layer is prepared on the surface of the substrate;

[0016] A support film is prepared on the surface of the sacrificial layer.

[0017] According to some embodiments of this application, the preparation of the substrate includes:

[0018] The substrate is prepared using silicon material.

[0019] According to some embodiments of this application, the preparation of the sacrificial layer on the surface of the substrate includes:

[0020] The sacrificial layer is prepared using phosphosilicate glass, borosilicate glass, or silicon oxide.

[0021] According to some embodiments of this application, etching the support film to form at least one accommodating channel includes:

[0022] Frontal exposure microscopy;

[0023] The support film is etched using a dry etching process to form the accommodating channel.

[0024] According to some embodiments of this application, a first support sheet is disposed on top of the support film, comprising:

[0025] The first support sheet is prepared using a light-transmitting material.

[0026] According to some embodiments of this application, a first support sheet is disposed on top of the support film, comprising:

[0027] The first support sheet is made of a material that can withstand at least 400°C.

[0028] According to some embodiments of this application, a support is prepared at the bottom of the support membrane, comprising:

[0029] Remove the first bottom silicon from the first substrate;

[0030] Etch the first substrate;

[0031] Fabricate the first support unit;

[0032] Remove the first support piece.

[0033] According to some embodiments of this application, a first support sheet is disposed on top of the support film, comprising:

[0034] A temporary bonding adhesion layer and a laser debonding layer are formed on top of the first support film;

[0035] The first support sheet is attached to the top of the laser debonding layer.

[0036] According to some embodiments of this application, removing the first bottom silicon in the first substrate includes:

[0037] The first bottom silicon in the first substrate is removed by dry etching or grinding.

[0038] According to some embodiments of this application, etching the first substrate includes:

[0039] The first silicon oxide layer and the first top silicon in the first substrate are etched to form a recessed region at the bottom of the first substrate to define the range of the first imaging window.

[0040] According to some embodiments of this application, the fabrication of the first support unit includes:

[0041] Within the range of the first imaging window, a plurality of first support units for supporting the first support film are prepared using the first top silicon and the first sacrificial layer in the first substrate.

[0042] According to some embodiments of this application, the fabrication of the first support unit includes:

[0043] The first top silicon in the first substrate is etched into the first sacrificial layer according to the grid shape to form a plurality of first support beams.

[0044] According to some embodiments of this application, the fabrication of the first support unit includes:

[0045] The first sacrificial layer is etched to the first support film according to the grid shape to form a plurality of second support beams, each of the second support beams and each of the first support beams corresponding one to one, and stacked to form each of the first support units.

[0046] According to some embodiments of this application, removing the first support sheet includes:

[0047] The first support sheet on top of the first support film is removed using a laser debonding process, and the temporary bonded adhesive layer is also removed.

[0048] According to some embodiments of this application, a support is prepared at the bottom of the support membrane, comprising:

[0049] Remove the second bottom silicon in the second substrate and remove the second sacrificial layer;

[0050] Prepare the first metal layer;

[0051] Prepare a titanium ring having a second metal layer and a second support sheet;

[0052] Bonding the first metal layer and the second metal layer;

[0053] Remove the first support piece and the second support piece.

[0054] According to some embodiments of this application, after etching the support film to form at least one accommodating channel, the method further includes:

[0055] A support structure is prepared on top of the second support membrane according to the grid shape to form a plurality of second support units for supporting the second support membrane.

[0056] According to some embodiments of this application, the step of preparing a support structure on top of the second support membrane according to the grid shape includes:

[0057] The support structure was fabricated using a lift-off process.

[0058] According to some embodiments of this application, removing the second bottom silicon in the second substrate and removing the second sacrificial layer includes:

[0059] The second substrate's second bottom silicon is removed by dry etching or polishing.

[0060] The second sacrificial layer was removed by wet etching.

[0061] According to some embodiments of this application, the preparation of the first metal layer includes:

[0062] A first metal layer is prepared at the bottom of the second support film.

[0063] According to some embodiments of this application, the preparation of the titanium ring having a second metal layer and a second support sheet includes:

[0064] A temporary bonding process is used to bond the titanium sheet to the second support sheet;

[0065] A second metal layer is deposited on the titanium sheet.

[0066] According to some embodiments of this application, the preparation of the titanium ring having a second metal layer and a second support sheet includes:

[0067] The second metal layer and the titanium sheet on the second support sheet are cut to form a ring structure corresponding to the shape of the first metal layer.

[0068] According to some embodiments of this application, cutting the second metal layer and the titanium sheet on the second support sheet includes:

[0069] The second metal layer and the titanium sheet are cut using a laser.

[0070] According to some embodiments of this application, bonding the first metal layer and the second metal layer includes:

[0071] The first metal layer and the second metal layer are bonded by eutectic bonding or metal hot pressing bonding.

[0072] According to some embodiments of this application, removing the first support sheet and the second support sheet includes:

[0073] The first support sheet and the second support sheet are removed using a laser debonding process.

[0074] According to some embodiments of this application, the preparation of the substrate and the preparation of a support film on top of the substrate includes:

[0075] Fabricate conductive material-filled areas;

[0076] Metal is deposited in the conductive material filling region.

[0077] According to some embodiments of this application, the fabrication of the conductive material filling region includes:

[0078] The third support film and the third substrate are etched down to the bottom of the silicon dioxide layer of the third substrate to form the conductive material filling region.

[0079] According to some embodiments of this application, depositing metal in the conductive material filling region includes:

[0080] A conductor is deposited in the conductive filling region, wherein the conductor is one of aluminum, titanium nitride, tungsten, or gold.

[0081] According to some embodiments of this application, after depositing a conductive material in the conductive material filling region, the process includes:

[0082] Annealing is performed to form an ohmic contact between the conductor and the third substrate.

[0083] According to some embodiments of this application, a support is prepared at the bottom of the support membrane, comprising:

[0084] A silicon oxide film is deposited on the bottom of the third substrate;

[0085] The third substrate is etched to form the third support beam by etching the third bottom silicon layer and the second silicon oxide layer.

[0086] Remove the first support sheet;

[0087] The second top silicon of the third substrate is etched to form the fourth support beam.

[0088] According to some embodiments of this application, depositing a silicon oxide film on the bottom of a third substrate includes:

[0089] Silicon oxide film was deposited using the ICP-VD process.

[0090] According to some embodiments of this application, removing the first support sheet includes:

[0091] The first support sheet is removed by acrylic cleaning or IPA cleaning.

[0092] According to a second aspect embodiment of this application, a grid structure for cryo-electron microscopy includes a support body, a support film, and an interlayer; the support film has at least one through-hole receiving channel, and the support film is made of silicon nitride, silicon carbide, or nickel-titanium alloy; the interlayer is located between the support film and the support body, and the hollow portions of the interlayer and the support body define an imaging window, with the receiving channel located within the area of ​​the imaging window.

[0093] The grid structure for cryo-electron microscopy according to the embodiments of this application has at least the following beneficial effects: The grid structure for cryo-electron microscopy applied to the first aspect of the embodiments of this application includes a support, a supporting film, and an interlayer. The interlayer is disposed between the supporting film and the support. On the one hand, the interlayer can provide support for the supporting film during the preparation of the supporting film, avoiding the decrease in vacuum degree of the grid structure caused by the pore making of the supporting film, and also avoiding damage and contamination of the film structure, ensuring the adsorption effect and flatness of the supporting film, and improving the yield. On the other hand, the supporting film can be combined with the metal support through the interlayer, or the interlayer can be used to deposit a conductive material, thereby improving the conductivity of the grid structure.

[0094] According to some embodiments of this application, the thickness of the support film is greater than or equal to 15 nm and less than or equal to 50 nm.

[0095] According to some embodiments of this application, the thickness of the support is greater than or equal to 20 μm and less than or equal to 100 μm.

[0096] According to some embodiments of this application, the grid structure for cryo-electron microscopy further includes a support structure, which is attached to the support film and is used to support the support film.

[0097] According to some embodiments of this application, the support structure is provided with a plurality of hollow portions, and the receiving channel is located within the range of each of the hollow portions.

[0098] According to some embodiments of this application, the support structure includes a plurality of support units, each of which is staggered within the range of the imaging window to form a grid structure, wherein the hollow portion of the grid structure defines each of the cutout portions.

[0099] According to some embodiments of this application, the thickness of the support structure is greater than or equal to 1 μm and less than or equal to 5 μm.

[0100] According to some embodiments of this application, the interlayer is made of phosphosilicate glass, borosilicate glass, or silicon oxide.

[0101] According to some embodiments of this application, the thickness of the interlayer is greater than or equal to 100 nm and less than or equal to 800 nm.

[0102] According to some embodiments of this application, the support unit includes a second support beam, each of the second support beams being attached to the bottom of the support membrane, and adjacent second support beams forming a second grid.

[0103] According to some embodiments of this application, each of the second support beams is formed by etching the interlayer.

[0104] According to some embodiments of this application, the support is made of silicon.

[0105] According to some embodiments of this application, the support unit further includes a first support beam, each first support beam being attached to the bottom of each second support beam, adjacent first support beams forming a first grid, each first grid and each second grid being connected in a one-to-one correspondence to form each of the hollow portions.

[0106] According to some embodiments of this application, each of the first support beams is formed by etching the support body.

[0107] According to some embodiments of this application, the thickness of the first support beam is less than the thickness of the support body.

[0108] According to some embodiments of this application, the interlayer is made of metal.

[0109] According to some embodiments of this application, the interlayer includes a first metal layer disposed at the bottom of the support film and a second metal layer disposed at the top of the support, wherein the first metal layer and the second metal layer are bonded together.

[0110] According to some embodiments of this application, the support is made of titanium.

[0111] According to some embodiments of this application, the support structure is made of titanium and is attached to the top of the support membrane.

[0112] According to some embodiments of this application, the width of the support unit is greater than or equal to 10 μm and less than or equal to 20 μm.

[0113] According to some embodiments of this application, the interlayer includes a top silicon layer and a silicon oxide layer stacked together, the silicon oxide layer being located between the top silicon layer and the support, and the interlayer having a deposition channel for depositing a conductor.

[0114] According to some embodiments of this application, the conductor is made of one of aluminum, titanium nitride, tungsten, or gold.

[0115] According to some embodiments of this application, the support unit includes a fourth support beam, each of the fourth support beams being attached to the bottom of the support membrane, and adjacent fourth support beams forming a fourth grid.

[0116] According to some embodiments of this application, the fourth grid is formed by etching the top silicon.

[0117] According to some embodiments of this application, the support unit further includes a third support beam, which is attached to the bottom of the silicon oxide layer. Adjacent third support beams form a third grid, and each third grid and each fourth grid are connected in a one-to-one correspondence to form a hollow portion.

[0118] According to some embodiments of this application, the third grid is formed by etching the support.

[0119] According to some embodiments of this application, the third grid is formed by etching the support.

[0120] Additional aspects and advantages of this application will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of this application. Attached Figure Description

[0121] The present application will be further illustrated below with reference to the accompanying drawings and embodiments. It should be noted that the embodiments illustrated in the following drawings are exemplary and are only used to explain the present application, and should not be construed as limiting the present application.

[0122] Figure 1 This is a schematic diagram of the first embodiment of the grid structure used in cryo-electron microscopy according to this application;

[0123] Figure 2 This is a schematic diagram of the second embodiment of the grid structure for cryo-electron microscopy in this application;

[0124] Figure 3 This is a schematic diagram of the third embodiment of the grid structure for cryo-electron microscopy in this application;

[0125] Figure 4 This is a schematic diagram of the substrate structure during the fabrication process of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0126] Figure 5 This is a schematic diagram of the sacrificial layer and supporting film during the preparation of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0127] Figure 6 This is a schematic diagram of the support film after etching during the fabrication process of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0128] Figure 7 This is a schematic diagram of the structure after the support sheet is attached to the top of the support membrane during the preparation of the first embodiment of the carrier structure for cryo-electron microscopy of this application;

[0129] Figure 8 This is a schematic diagram of the structure after removing the bottom silicon of the substrate during the fabrication process of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0130] Figure 9 This is a schematic diagram of the structure after etching the silicon oxide layer and top silicon in the substrate during the fabrication process of the first embodiment of the carrier structure used in cryo-electron microscopy of this application;

[0131] Figure 10 This is a schematic diagram of the first support beam formed by etching during the fabrication process of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0132] Figure 11 This is a schematic diagram of the second support beam etched during the fabrication process of the first embodiment of the carrier structure for cryo-electron microscopy in this application;

[0133] Figure 12 This is a schematic diagram of the structure after removing the top support sheet of the support film during the preparation process of the first embodiment of the carrier structure for cryo-electron microscopy of this application;

[0134] Figure 13 This is a schematic diagram of the substrate structure during the fabrication process of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0135] Figure 14 This is a schematic diagram of the support film after etching during the fabrication process of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0136] Figure 15 This is a schematic diagram of the structure after the support structure is prepared during the preparation process of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0137] Figure 16 This is a schematic diagram of the structure after the support sheet is attached to the support film and the top of the support structure during the preparation of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0138] Figure 17 This is a schematic diagram of the structure after removing the underlying silicon and sacrificial layer and preparing the first metal layer during the preparation process of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0139] Figure 18 This is a schematic diagram of the structure after the titanium sheet, support sheet, and second metal layer are combined during the preparation of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0140] Figure 19 This is a schematic diagram of the titanium ring structure during the preparation of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0141] Figure 20 This is a schematic diagram of the structure after the first metal layer and the second metal layer are combined during the preparation of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0142] Figure 21 This is a schematic diagram of the structure after removing the support sheet during the preparation process of the second embodiment of the carrier structure for cryo-electron microscopy in this application;

[0143] Figure 22 This is a schematic diagram of the first scenario of the third embodiment of the grid structure used in cryo-electron microscopy in this application;

[0144] Figure 23 This is an exploded view of the structure in the second case of the third embodiment of the grid structure used for cryo-electron microscopy in this application;

[0145] Figure 24 This is a schematic diagram of the structure after a support film is prepared on top of the substrate during the fabrication process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0146] Figure 25 This is a schematic diagram of the structure after the conductive material filling region is fabricated during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0147] Figure 26This is a schematic diagram of the structure after metal deposition in the conductive material filling area during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0148] Figure 27 This is a schematic diagram of the structure after etching the support film during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0149] Figure 28 This is a schematic diagram of the structure after the first support sheet is placed on top of the support membrane during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy of this application;

[0150] Figure 29 This is a schematic diagram of the substrate after a silicon oxide film is deposited on the bottom of the substrate during the fabrication process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0151] Figure 30 This is a schematic diagram of the structure after the third support beam is formed during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0152] Figure 31 This is a schematic diagram of the structure after removing the first support sheet during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application;

[0153] Figure 32 This is a schematic diagram of the structure after the fourth support beam is formed during the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy in this application.

[0154] Figure label:

[0155] 101a. First support; 101b. Second support; 101c. Third support; 102a. First support film; 102b. Second support film; 102c. Third support film; 1021a. First accommodating channel; 1021b. Second accommodating channel; 1021c. Third accommodating channel; 103a. First interlayer; 103b. Second interlayer; 103c. Third interlayer; 104. First metal layer; 105. Second metal layer; 106a. First imaging window; 106b. Second imaging window; 106c. Third imaging window;

[0156] 201a. First support unit; 201b. Second support unit; 201c. Third support unit; 202. First support beam; 203. Second support beam; 204. Third support beam; 205. Fourth support beam;

[0157] 301a. First substrate; 301b. Second substrate; 301c. Third substrate; 302a. First bottom silicon; 302b. Second bottom silicon; 302c. Third bottom silicon; 303a. First silicon oxide layer; 303b. Second silicon oxide layer; 304a. First top silicon; 304b. Second top silicon; 305a. First sacrificial layer; 305b. Second sacrificial layer; 306. First support sheet; 307. Laser debonding layer; 308. Temporary bonding adhesive layer;

[0158] 401. Second support sheet; 402. Conductive material filling area; 403. Conductor. Detailed Implementation

[0159] The embodiments of this application are described in detail below with reference to the accompanying drawings, wherein the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are exemplary and are only used to explain this application, and should not be construed as limiting this application.

[0160] In the description of this application, it should be understood that the terms "center", "middle", "longitudinal", "transverse", "length", "width", "thickness", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", "axial", "radial", "circumferential", etc., indicating the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings, are only for the convenience of describing this application and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this application.

[0161] In the description of this application, "several" means one or more, "multiple" means two or more, "greater than," "less than," and "exceeding" are understood to exclude the stated number, while "above," "below," and "within" are understood to include the stated number. The use of "first" and "second" in the description is merely for distinguishing technical features and should not be construed as indicating or implying relative importance, or implicitly indicating the number of indicated technical features, or implicitly indicating the order of the indicated technical features.

[0162] In the description of this application, unless otherwise expressly specified and limited, the terms "set up," "install," "connect," and "link" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal connection of two components. Those skilled in the art can understand the specific meaning of the above terms in this application based on the specific circumstances.

[0163] In the description of this application, the use of terms such as "one embodiment," "some embodiments," "an example," "some instances," "some embodiments," "illustrative embodiment," "example," "specific example," and "some examples" indicates that the specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of this application. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0164] like Figure 1 , Figure 2 and Figure 3 As shown in the embodiments of this application, a grid structure for cryo-electron microscopy is provided. The grid structure for cryo-electron microscopy includes a support body, a support film, and an interlayer. In the first embodiment, the support body is a first support body 101a; in the second embodiment, it is a second support body 101b; and in the third embodiment, it is a third support body 101c. In the first embodiment, the support film is a first support film 102a; in the second embodiment, it is a second support film 102b; and in the third embodiment, it is a third support film 102c. In the first embodiment, the interlayer is a first interlayer 103a; in the second embodiment, it is a second interlayer 103b; and in the third embodiment, it is a third interlayer 103c.

[0165] The support membrane holds the sample solution, and the support body supports the membrane at its bottom. The flat bottom surface of the support body ensures the stable placement of the grid structure on the cryo-electron microscope, facilitating sample observation by the operator.

[0166] Furthermore, the interlayer is located between the support and the supporting membrane. The interlayers in this application are made of different materials and have different structural forms. In the first embodiment, the first interlayer 103a can be made of an inorganic non-metallic material. Existing processing techniques first prepare the support and its imaging window, and then fabricate the accommodating channel from the top surface of the supporting membrane. This processing sequence can lead to vacuum leakage problems in some stages. Specifically, the accommodating channel is the first accommodating channel 1021a in the first embodiment, the second accommodating channel 1021b in the second embodiment, and the third accommodating channel 1021c in the third embodiment.

[0167] In the fabrication of the first support film 102a in the first embodiment of this application, the first interlayer 103a and the first support 101a are located at the bottom of the first support film 102a. This avoids the problem of vacuum level reduction caused by pore formation in the first support film 102a, and also prevents damage and contamination to the first support film 102a, ensuring the adsorption effect of the carrier structure, i.e., ensuring the yield rate. At the same time, the first interlayer 103a can provide support for the first support film 102a. Under the support, the size, shape, and characteristics of the carrier structure can be precisely controlled, including the size, spacing, and distribution of pores on the first support film 102a, as well as the overall thickness and geometry of the carrier structure.

[0168] On the other hand, in the second embodiment, the second substrate 301b is located at the bottom of the second support film 102b, thereby avoiding the problem of vacuum level reduction caused by forming holes in the second support film 102b, and also avoiding damage and contamination to the second support film 102b. Simultaneously, after the second interlayer 103b is formed at the bottom of the second support film 102b, since the second interlayer 103b is made of metal, it facilitates the bonding of the metal second support 101b to the second support film 102b through the second interlayer 103b, thereby improving the conductivity of the carrier structure.

[0169] In addition, in the third embodiment, the interlayer is the second silicon oxide layer 303b and the second top silicon layer 304b located at the bottom of the third support film 102c, and the third support 101c is also located at the bottom of the third support film 102c. This avoids the problem of vacuum reduction caused by pore formation in the third support film 102c, and also avoids damage and contamination to the second support film 102b. Simultaneously, the interlayer also provides channels for depositing conductive materials to reduce the resistivity of the grid structure, thereby further improving the conductivity of the grid structure. Furthermore, the significant improvement in conductivity can reduce the BIM phenomenon caused by charging during sample observation, greatly improve imaging quality, and provide strong support for high-precision cryo-electron microscopy observation.

[0170] The fabrication method for the carrier structure in this application follows the steps of MEMS fabrication technology, enabling mass production and reducing the risks of contamination and damage during sample preparation and imaging. It allows for the rapid manufacture of a large number of carrier structures to meet practical application needs. Precise control of the size, shape, and features of the carrier structure ensures high consistency in mass-produced carrier structures, thereby improving the quality of the carrier products.

[0171] In addition, the support is located at the edge of the support film, and the thickness of the support is greater than the thickness of the support film, so that the carrier structure is formed as a structure that is thin in the middle and thick at the edge. The larger edge thickness is beneficial to improving the overall strength of the carrier. Typically, the minimum thickness of continuous film deposition in CVD can be 15nm. The thickness of the support film in this application can be set to 15nm to 50nm. Specifically, the thickness of the support film in this application is set to 20nm to 50nm.

[0172] It is worth noting that the surface of the support membrane is usually provided with support beams that provide support and ensure the strength of the support membrane. Understandably, a thicker support beam provides stronger support; however, a thicker support beam can cause shadows to obstruct the observation area during electron microscopy. Therefore, in the third embodiment of this application, the support beams include multiple layers, namely a third support beam 204 and a fourth support beam 205. The placement of the third support beam 204 significantly enhances the strength of the carrier structure, allowing for a substantial reduction in the thickness of the fourth support beam 205, which is closer to the third support membrane 102c, from the previous 15μm-50μm to 2μm-5μm. This essentially eliminates the obstruction of the observation area by shadows during electron microscopy, significantly improving the clarity and integrity of the observation field.

[0173] Because the supporting membrane is thinner, it facilitates obtaining a thinner and more uniform ice layer during subsequent biological sample preparation, ensuring the imaging effect of the grid. However, the CTE coefficient can cause the supporting membrane to bend when it is attached to the support sheet, and processing stress can also cause bending after the support sheet detaches from the supporting membrane. The supporting membrane surface of this application is provided with support units. Under the support of these units, the degree of bending / overall span of the supporting membrane in this application is numerically smaller than that of the prior art, resulting in a flatter supporting membrane, which is beneficial for grid imaging.

[0174] In some examples, the support membrane has at least one accommodating channel, which is formed by pore fabrication. The accommodating channel extends through the top and bottom of the support membrane and serves to contain the sample solution, ensuring sample adhesion to the support membrane. After the sample is attached to the support membrane, the electron beam emitted by the electron microscope can penetrate the sample, thereby completing the image formation.

[0175] Specifically, the accommodating channel is located in the middle of the support membrane, that is, the sample is attached to the middle of the support membrane, while the edge of the support membrane is used to set the support body.

[0176] Furthermore, both the interlayer and the support are formed as hollow annular structures. The interlayer is located between the support membrane and the support, and the support membrane is connected to the support through the interlayer. The radial dimensions of the support membrane and the support are approximately the same, allowing the hollow portions of the interlayer and the support to jointly define the imaging window of the carrier structure. In the first embodiment, the imaging window is a first imaging window 106a; in the second embodiment, it is a second imaging window 106b; and in the third embodiment, it is a third imaging window 106c. Each accommodating channel needs to be located within the imaging window to prevent the interlayer and the support from obstructing the channels, facilitating the smooth passage of the electron beam through the sample and support membrane, carrying the sample's structural information to the detector, thereby forming a clear image.

[0177] In some examples, the support membrane is made of silicon nitride, silicon carbide, or nickel-titanium alloy.

[0178] The silicon nitride support film exhibits excellent chemical inertness, resisting chemical reactions when irradiated by an electron beam or in contact with various biological samples or chemical reagents. Furthermore, the silicon nitride support film possesses a certain degree of mechanical strength, making it resistant to breakage or deformation. Simultaneously, the silicon nitride support film allows for electron beam penetration, enabling imaging and avoiding imaging tremors. Before observation using an electron microscope, sample alignment is required. Specifically, images of the support film and substrate are taken, and the images are adjusted in real-time. Diffraction rings generated by the support film are observed using diffraction modes. When using silicon nitride, diffraction rings can be generated in diffraction modes. In the first embodiment, the substrate is the first substrate 301a; in the second embodiment, it is the second substrate 301b; and in the third embodiment, it is the third substrate 301c.

[0179] Furthermore, silicon carbide support films have high hardness, resulting in support films with good wear resistance. They can maintain stable physical and chemical properties even at high temperatures and are less likely to release impurities that contaminate the sample during electron microscopy experiments.

[0180] In addition, the nickel-titanium alloy support film has shape memory effect and superelasticity, which can better fit the sample, and the nickel-titanium alloy support film has a certain degree of conductivity.

[0181] Staff can choose silicon nitride, silicon carbide, or nickel-titanium alloy materials to make the support membrane according to actual needs.

[0182] In some examples, the thickness of the support film is greater than or equal to 15 nm and less than or equal to 50 nm, and the thinner support film can meet the needs of Vitrobot's fabrication.

[0183] In the Vitrobot fabrication process, the mesh is first held upright by tweezers. Then, a drop of liquid is added to the mesh, and two sheets of filter paper are placed on either side of the mesh to hold it in place. Excess liquid is then absorbed. In this scenario, the mesh itself needs to be very thin to ensure the liquid on its surface can be completely absorbed. However, existing meshes are approximately 200μm to 300μm thick, which is unsuitable for this operation.

[0184] Furthermore, the relatively small overall thickness of the carrier mesh ensures the penetration effect of the electron beam and guarantees the clarity of the image.

[0185] Furthermore, the thinner mesh allows the sample to be arranged in a single layer within the support film, minimizing overlap of sample particles in the height direction. This avoids overlap during imaging and ensures that all sample particles are on the same focal plane, facilitating subsequent data processing.

[0186] In some examples, the thickness of the support is greater than or equal to 20 μm and less than or equal to 100 μm. This relatively large thickness relative to the support membrane ensures the stability of the support membrane.

[0187] In some examples, the grid structure used for cryo-electron microscopy also includes a support structure that is attached to the support membrane. Understandably, the strength of the support structure is greater than the strength of the support membrane, thus providing support for the support membrane, ensuring its flatness, and preventing warping.

[0188] The support structure can be located at the bottom of the support membrane, such as the support structure composed of the first support unit 201a in the first embodiment or the third support unit 201c in the third embodiment; or it can be located at the top of the support membrane, such as the support structure composed of the second support unit 201b in the second embodiment. It is worth noting that when the support structure is located at the top of the support membrane, the support structure needs to avoid the receiving channel of the support membrane to ensure that the sample solution can enter the receiving channel.

[0189] In some examples, the support structure has several hollow sections, and the receiving channel is located within the hollow section. On the one hand, this ensures that the sample solution can smoothly enter the receiving channel; on the other hand, it ensures that the electron beam carrying sample information can reach the detector of the cryo-electron microscope.

[0190] It is understandable that the portion of the support membrane located outside the imaging window is supported by the support body, eliminating the need for a separate support structure. Therefore, the support structure is located within the imaging window.

[0191] In some examples, the support structure comprises several support units, which are staggered and form a grid shape within the imaging window. This grid-shaped support structure, while ensuring support, avoids the accommodating channels on the support membrane; the hollow portion of the grid-shaped support structure is the aforementioned perforated section.

[0192] In some examples, the support structure has a relatively small thickness, specifically, greater than or equal to 1 μm and less than or equal to 5 μm. If the support structure is too thick, when the sample solution enters the containment channel and forms a liquid film according to the grid shape of the support structure, the liquid film will have a large thickness at the angular position where the support unit and the support film meet, which is not conducive to imaging observation. However, the support structure of this application has a smaller thickness, which is conducive to maintaining a relatively uniform thickness of the liquid film at various positions, thereby improving the imaging accuracy.

[0193] In the first embodiment, as Figure 1 As shown, the first interlayer 103a serves as the first sacrificial layer 305a. On one hand, the first interlayer 103a, as the first sacrificial layer 305a, can provide support during the preparation of the first support film 102a. On the other hand, the first interlayer 103a is also used to form the first support unit 201a located at the bottom of the first support film 102a. Compared with the material of the first support film 102a, the material of the first sacrificial layer 305a is harder. Therefore, the first support unit 201a formed by the first sacrificial layer 305a can provide support for the first support film 102a.

[0194] The first interlayer 103a is made of inorganic non-metallic material, specifically, the first interlayer 103a is made of phosphosilicate glass, borosilicate glass or silicon oxide.

[0195] When the first interlayer 103a is made of phosphosilicate glass, it has advantages such as low melting point, easy removal, and good surface flatness; when the first interlayer 103a is made of borosilicate glass, it has advantages such as flexible adjustment performance and strong stress buffering ability; when the first interlayer 103a is made of silicon oxide, it has advantages such as high chemical stability and good insulation performance.

[0196] Staff can choose the material of the first interlayer 103a as the first sacrificial layer 305a according to actual needs.

[0197] In the first embodiment, the thickness of the first interlayer 103a is greater than or equal to 100 nm and less than or equal to 800 nm. Compared with the first support film 102a, the first interlayer 103a has a larger thickness, which can provide sufficient support for the first support film 102a during the preparation of the first support film 102a, thereby ensuring the quality of the prepared first support film 102a.

[0198] In the first embodiment, the first support unit 201a includes a second support beam 203, which is attached to the bottom of the first support membrane 102a to provide support for the first support membrane 102a. Depending on the arrangement of the first support unit 201a, each second support beam 203 is also formed in a grid shape, meaning that adjacent second support beams can form a second grid, and each second grid is used to form a cutout portion.

[0199] It is worth noting that each of the second support beams 203 and the first interlayer 103a are integral, that is, each of the second support beams 203 and the first sacrificial layer 305a are integral. Specifically, within the range of the first imaging window 106a, each of the second support beams 203 is formed by etching the first interlayer 103a.

[0200] In the first embodiment, the first substrate 301a used in the fabrication process of the carrier structure is made of silicon. The silicon first substrate 301a is retained in the finished carrier structure and forms the first support 101a.

[0201] Furthermore, the first support unit 201a also includes a first support beam 202, which is attached to the bottom of the second support beam 203 to further provide support for the first support membrane 102a. It can be understood that, according to the arrangement of the first support unit 201a, each first support beam 202 is formed in a grid shape, and the grid shape formed by each first support beam 202 corresponds to the grid shape formed by each second support beam 203, facilitating a one-to-one correspondence between each first support beam 202 and each second support beam 203, and their combination to form the first support unit 201a.

[0202] Among them, adjacent first support beams 202 can form a first grid, and each first grid is connected to each second grid in a one-to-one correspondence, thereby forming the hollow part of the support structure.

[0203] It is worth noting that each of the first support beams 202 and the first support body 101a are an integral whole. Specifically, within the range of the first imaging window 106a, the first support body 101a forms each of the first support beams 202 by etching.

[0204] In the first embodiment, since there is no first accommodating channel 1021a within the range of the first support 101a, the thickness of the support is relatively large to ensure the support stability of the first support membrane 102a. The first support unit 201a needs to have a smaller thickness to avoid a significant increase in the thickness of the liquid film formed by the sample solution at the interface between the first support unit 201a and the first support membrane 102a. Therefore, the first support beam 202 forming the first support unit 201a also needs to have a smaller thickness, and the thickness of the first support beam 202 is less than the thickness of the first support 101a.

[0205] In the second embodiment, as Figure 2 As shown, the second interlayer 103b is made of metal, which facilitates the bonding of the metal second support 101b to the second support film 102b through the second interlayer 103b, thereby improving the conductivity of the carrier structure.

[0206] In the second embodiment, the second support 101b is made of titanium, which has a certain degree of conductivity to improve the overall conductivity of the carrier structure. However, the titanium second support 101b is difficult to bond to the second support membrane 102b, and a second interlayer 103b is required to ensure the connection between the second support 101b and the second support membrane 102b. Therefore, the second interlayer 103b in the second embodiment is not the second sacrificial layer 305b, but a structure used to ensure a stable connection between the second support 101b and the second support membrane 102b.

[0207] Furthermore, the second interlayer 103b includes a first metal layer 104 and a second metal layer 105. The first metal layer 104 is fixedly disposed at the bottom of the second support film 102b, and the second metal layer 105 is fixedly disposed at the top of the titanium material second support body 101b. The first metal layer 104 and the second metal layer 105 are formed into a whole through a bonding process, thereby ensuring that the second support film 102b and the second support body 101b are connected into a whole.

[0208] Specifically, the bonding process between the first metal layer 104 and the second metal layer 105 adopts a permanent bonding process.

[0209] In the second embodiment, a support structure is fabricated on top of the second support film 102b, and the support structure is made of titanium. Since both the support structure at the top of the second support film 102b and the second support body 101b at the bottom of the second support film 102b are made of titanium, the overall conductivity of the carrier structure can be improved.

[0210] Among them, a highly conductive carrier structure can reduce the charge accumulation induced by the electron beam and improve the transmission efficiency of electronic signals, thereby ensuring imaging quality.

[0211] It is understood that the support structure of the second embodiment is attached to the top of the second support film 102b, thereby providing support for the second support film 102b.

[0212] In the second embodiment, the second support unit 201b is also formed in a grid shape, and adjacent second support units 201b form a corresponding grid, where the grid forms a hollow part.

[0213] The width of the second support unit 201b of the support structure is greater than or equal to 10μm and less than or equal to 20μm. The specific width of the second support unit 201b can be set according to the distribution of the second accommodating channel 1021b to ensure that each second accommodating channel 1021b is within the range of the hollow part, so as to facilitate the sample solution to enter the second accommodating channel 1021b.

[0214] In the third embodiment, as Figure 3 and Figure 24 As shown, the third substrate 301c includes a second top silicon 304b, a third bottom silicon 302c, and a second silicon oxide layer 303b. The third bottom silicon 302c is used to form the third support 101c, while the second top silicon 304b and the second silicon oxide layer 303b are used to form an interlayer, that is, the interlayer includes the second top silicon 304b and the second silicon oxide layer 303b stacked together.

[0215] Among them, such as Figure 25 As shown, the interlayer has deposition channels that can form conductive material filling regions 402 on the carrier structure. Specifically, the second top silicon 304b and the second silicon oxide layer 303b both have corresponding pore structures. The pore structures of the second top silicon 304b and the second silicon oxide layer 303b correspond to each other in position. The pore structures of the second top silicon 304b and the second silicon oxide layer 303b together define the deposition channels, thereby forming the conductive material filling regions 402. At the same time, the surface of the third support film 102c also has corresponding pore structures. The pore structures of the third support film 102c correspond to the deposition channels in position, facilitating the entry of the conductor 403 from the surface of the third support film 102c into the deposition channels, thereby completing the deposition process of the conductor 403 in the conductive material filling regions 402.

[0216] Furthermore, compared to the first and second embodiments, the third embodiment eliminates the sacrificial layer, fundamentally reducing the overall resistivity of the carrier structure. Simultaneously, depositing the conductor 403 in the deposition channels further reduces the resistivity of the carrier structure, significantly improving the conductivity of the sample.

[0217] In the third embodiment, the conductor 403 is made of one of aluminum, titanium nitride, tungsten, or gold. To ensure the electrical connection between the conductor 403 and other structures in the carrier network, rapid annealing is required after the conductor 403 is deposited, so that the conductor 403 forms an ohmic contact.

[0218] In the third embodiment, the third support unit 201c includes a fourth support beam 205, which is attached to the bottom of the third support membrane 102c to provide support for the third support membrane 102c. Each fourth support beam 205 is formed in a grid shape, that is, adjacent fourth support beams can form a fourth grid, and each fourth grid is used to form a cutout.

[0219] It is worth noting that each of the fourth support beams 205 is integral with the interlayer; specifically, each of the fourth support beams 205 is integral with the second top silicon 304b. Specifically, within the range of the third imaging window 106c, each of the fourth support beams 205 is formed by etching the second top silicon 304b.

[0220] In the third embodiment, the third support unit 201c further includes a third support beam 204, which is located at the bottom of the fourth support beam 205. It is understood that both the third and fourth support beams 204 and 205 can provide support for the third support membrane 102c. The inclusion of the third support beam 204 significantly enhances the overall strength of the carrier structure, allowing for a substantial reduction in the thickness of the fourth support beam 205, from 15μm to 50μm in conventional designs to 2μm to 5μm. This reduced height of the fourth support beam 205 eliminates the obstruction of the observation area by shadows during electron microscopy, significantly improving the clarity and integrity of the observation field.

[0221] It is understood that the third support beam 204 is formed in a grid shape, and the grid shape formed by each third support beam 204 corresponds to the grid shape formed by each fourth support beam 205, so that each third support beam 204 and each fourth support beam 205 can be matched one-to-one and combined to form the third support unit 201c.

[0222] The adjacent third support beams 204 can form a third grid, and each third grid is connected to each fourth grid in a one-to-one correspondence, thus forming the hollow part of the support structure. At the same time, additional support beams and other support structures can be set in the hollow part according to actual needs, so as to ensure the strength of the net structure.

[0223] It is worth noting that each of the third support beams 204 and the third support body 101c are an integral unit. Specifically, within the range of the third imaging window 106c, the third support body 101c forms each of the third support beams 204 by etching.

[0224] in addition, Figure 22 and Figure 23 Two specific scenarios of the third embodiment are given. Figure 22 and Figure 23 The difference is that, Figure 22 The grid formed by the supporting beams in the illustrated case is hexagonal in shape, while Figure 23 The grid shape formed by the supporting beams in the example shown is rectangular.

[0225] Based on the above-described grid structure for cryo-electron microscopy, various embodiments of the fabrication method of the grid structure of this application are presented below.

[0226] The method for preparing this grid structure can be applied to the aforementioned grid structure used in cryo-electron microscopy. The method for preparing this grid structure includes, but is not limited to, the following steps:

[0227] Reference Figure 4 and Figure 5 Step S1: Prepare a substrate and prepare a support film on top of the substrate;

[0228] Reference Figure 6 Step S2: Etch the support film to form at least one accommodating channel;

[0229] Reference Figure 7 Step S3: A first support sheet 306 is disposed on the top of the support membrane;

[0230] Reference Figures 8 to 11 Step S4: Prepare a support at the bottom of the support membrane.

[0231] In some examples, the substrate is made using a standard SOI wafer, and the support film can be fabricated using processes such as LPCVD, ALD / PE-ALD / ICPCVD, etc. It is worth noting that PECVD cannot be used for the support film because PECVD typically produces films with thicknesses ranging from tens of nanometers to micrometers, which is insufficient for ultra-thin applications. Furthermore, PECVD requires high temperature and vacuum conditions, while the substrate material may oxidize or deform at high temperatures.

[0232] Furthermore, after the substrate and support film are prepared, the support film needs to be etched to form a accommodating channel. The etching process needs to penetrate the support film to create a accommodating channel that runs through the support film. The position of the accommodating channel is used to hold the sample, so that the electron beam emitted by the electron microscope can pass through the sample within the accommodating channel.

[0233] Meanwhile, a first support sheet 306 needs to be set on the top of the support membrane. The first support sheet 306 can be bonded to the top of the support membrane using a process such as temporary bonding. The first support sheet 306 can provide support for the support membrane on the top of the support membrane, which facilitates the subsequent process of preparing a support body at the bottom of the support membrane and avoids affecting the structure of the support membrane.

[0234] In addition, the support at the bottom of the support membrane can be made using subtractive processes such as etching, which can improve the strength of the carrier structure.

[0235] Additionally, step S1: preparing a substrate and preparing a support film on top of the substrate may include, but is not limited to, the following steps:

[0236] Reference Figure 5 Step S11: Prepare a sacrificial layer on the surface of the substrate.

[0237] Step S12: Prepare a support film on the surface of the sacrificial layer.

[0238] In some examples, the fabrication of the grid structure for cryo-electron microscopy first involves preparing a substrate and a support film.

[0239] In both the first and second embodiments, a sacrificial layer is provided. This sacrificial layer needs to be easily removable, capable of being removed in a relatively gentle manner during the experiment, without damaging the sample or affecting the performance of the grid. Furthermore, the sacrificial layer must be completely removed without any residue.

[0240] Furthermore, the sacrificial layer needs to have good adhesion, enabling it to adhere tightly and uniformly between the substrate and the supporting film to ensure the flatness of the supporting film and the imaging effect. In addition, the sacrificial layer also needs to be easy to prepare for subsequent processing.

[0241] Specifically, the sacrificial layer can be made of LPCVD-TEOS silicon oxide, LPCVD-BSPG silicon oxide, or PECVD-BSPG annealed silicon oxide.

[0242] It is worth noting that in the fabrication process of the net structure in the first embodiment, it is not necessary to completely remove the first sacrificial layer 305a; the first sacrificial layer 305a exists in the finished net structure as the second support beam 203 and the first interlayer 103a. However, in the fabrication process of the net structure in the second embodiment, it is necessary to completely remove the second sacrificial layer 305b.

[0243] In addition, the support film is etched to form accommodating channels; specifically, this etching is dry etching. Dry etching refers to a process that uses physical or chemical processes in a dry environment, such as plasma ion beams, to remove specific areas from the material surface. Dry etching can meet the high precision requirements of cryo-electron microscopy for the hole patterns of the support film and can form hole structures with vertical sidewalls, avoiding problems such as electron beam scattering caused by sidewall tilt.

[0244] Specifically, to facilitate the support of the support film during subsequent substrate processing, the first support sheet 306 is attached to the top of the support film to prevent the substrate processing from affecting the support film.

[0245] Additionally, refer to Figure 4 , Figure 13 and Figure 24 Step S1: Prepare a substrate and prepare a support film on top of the substrate, which may include, but is not limited to, the following steps:

[0246] Step S13: Prepare the substrate using silicon material.

[0247] In some examples, silicon substrates exhibit high hardness and strength, enabling them to withstand various external forces that may occur during cryo-electron microscopy experiments. Simultaneously, silicon has a relatively low coefficient of thermal expansion, allowing it to maintain dimensional stability better in environments with significant temperature variations. Furthermore, silicon possesses excellent electrical properties and chemical stability.

[0248] Specifically, in the first embodiment, the substrate includes a bottom silicon layer, a top silicon layer, and a silicon oxide layer, with the silicon oxide layer disposed between the bottom silicon layer and the top silicon layer. In the second embodiment, the second substrate 301b only contains a second bottom silicon layer 302b. Specifically, the bottom silicon layer is a first bottom silicon layer 302a in the first embodiment, a second bottom silicon layer 302b in the second embodiment, and a third bottom silicon layer 302c in the third embodiment; the silicon oxide layer is a first silicon oxide layer 303a in the first embodiment and a second silicon oxide layer 303b in the third embodiment; and the top silicon layer is a first top silicon layer 304a in the first embodiment and a second top silicon layer 304b in the third embodiment.

[0249] The substrate adopts an all-silicon structure, which can achieve a high flatness (below 1μm), meet the requirements of electron microscopy observation, and achieve the effect of not needing secondary focusing.

[0250] Additionally, refer to Figure 5 and Figure 13 Step S11: Prepare a sacrificial layer on the surface of the substrate, which may include, but is not limited to, the following steps:

[0251] Step S111: Prepare the sacrificial layer using phosphosilicate glass, borosilicate glass, or silicon oxide.

[0252] Additionally, refer to Figure 6 and Figure 14 Step S2: Etching the support film to form at least one accommodating channel may include, but is not limited to, the following steps:

[0253] Step S21: Frontal exposure microscopy;

[0254] Step S22: Dry etching is used to form the accommodating channel.

[0255] In some examples, it is understandable that positive exposure helps to obtain clear and accurate image information.

[0256] Since the support film is prepared on the surface of the interlayer, when dry etching is performed according to the position and size of the accommodating channel, when the etching reaches the surface of the interlayer, it means that the current accommodating channel has penetrated through the support film.

[0257] Additionally, step S3: providing a first support sheet 306 on top of the support film may include, but is not limited to, the following steps:

[0258] Step S31: The first support sheet 306 is prepared using a light-transmitting material.

[0259] In some examples, the first support sheet 306 can be bonded to the top of the support film, and then the first support sheet 306 can be removed by laser debonding.

[0260] Furthermore, laser debonding has unique advantages for transparent materials such as glass and quartz. Lasers can penetrate these materials, focusing on the bonding interface and generating localized high energy density to induce physical or chemical changes in the bonded materials, thereby achieving bond separation.

[0261] Specifically, the first support sheet 306 is made of glass. The debonding of the first support sheet 306 and the support film can be achieved by using the high energy of the laser to precisely destroy the bonding layer.

[0262] Additionally, step S3: providing a first support sheet 306 on top of the support film may include, but is not limited to, the following steps:

[0263] Step S32: The first support sheet 306 is prepared using a material that can withstand at least 400°C.

[0264] In some examples, the process for preparing the first support sheet 306 includes homogenization, baking, and bonding, or homogenization, pre-baking, and bonding. Specifically, when using the baking process, the operating temperature of the bonding heater is 210°C, the heater pin rising temperature is 180°C, the heater idle temperature is 140°C, the homogenization delay is 600s, the bonding pressure output is 150N, and the platen bonding time is 600s. When using the pre-baking process, the operating temperature of the bonding heater is adjusted to 230°C.

[0265] The first support sheet 306, made of high-temperature resistant material, is easy to remove using laser debonding. Laser debonding technology can precisely manipulate the bonding interface at relatively high temperatures. By using a high-energy-density laser beam to act on the bonding site, the bonded material is decomposed or separated without causing excessive damage to the material itself.

[0266] Additionally, refer to Figures 4 to 17 In the fabrication process of the grid structure for cryo-electron microscopy in the first embodiment, step S4: fabricating a support at the bottom of the support film may include, but is not limited to, the following steps:

[0267] Step S41: Remove the first bottom silicon 302a from the first substrate 301a;

[0268] Step S42: Etch the first substrate 301a;

[0269] Step S43: Fabricate the first support unit 201a;

[0270] Step S44: Remove the first support piece 306.

[0271] In some examples, the removal of the first bottom silicon 302a from the first substrate 301a can be achieved using processes such as etching and polishing. Specifically, the etching here can be performed using ICP dry etching. Removing the first bottom silicon 302a can thin the first substrate 301a and expose the first silicon oxide layer 303a.

[0272] Furthermore, after removing the first bottom silicon 302a, the first substrate 301a needs to be etched to form the first imaging window 106a at the bottom of the carrier structure, and to form a silicon ring of about 50 μm on the first substrate 301a.

[0273] In addition, after forming the first imaging window 106a, a first support unit 201a needs to be fabricated. The first support unit 201a can support the support film, thereby improving the strength of the carrier structure and preventing deformation of the support film. Specifically, the first support unit 201a is fabricated by etching a first top silicon layer 304a and a first sacrificial layer 305a. Specifically, the top silicon layer of the substrate 301a can be etched using deep silicon etching (DRIE) or plasma etching (ICP), while the etching of the first sacrificial layer 305a can be etched using VHF or BOE etching.

[0274] Furthermore, after the first support unit 201a is prepared, the first support sheet 306 needs to be removed to complete the preparation process of the carrier structure of the first embodiment.

[0275] Additionally, refer to Figure 7 Step S3: A first support sheet 306 is disposed on top of the support film, which may include, but is not limited to, the following steps:

[0276] Step S33: A temporary bonding adhesion layer 308 and a laser debonding layer 307 are formed on the top of the first support film 102a;

[0277] Step S34: The first support sheet 306 is attached to the top of the laser debonding layer 307.

[0278] In some examples, a temporary bonding adhesive layer 308 is attached to the top of the first support film 102a, and a laser debonding layer 307 is located between the temporary bonding adhesive layer 308 and the first support sheet 306. When the laser irradiates the laser debonding layer 307, the first support sheet 306 can be separated from the first support film 102a, thereby facilitating the removal of the first support sheet 306.

[0279] The bonding process includes homogenization, baking, and bonding, or homogenization, pre-baking, and bonding. When using the baking process, the bonding heater's operating temperature is 210℃, the heater pin rising temperature is 180℃, the heater idle temperature is 140℃, the homogenization delay is 600s, the bonding pressure output is 150N, and the platen bonding time is 600s. When using the pre-baking process, the bonding heater's operating temperature is adjusted to 230℃. Additionally, refer to... Figure 8 Step S41: Removing the first bottom silicon 302a from the first substrate 301a may include, but is not limited to, the following steps:

[0280] Step S411: Remove the first bottom silicon 302a in the first substrate 301a by dry etching or grinding.

[0281] In some examples, the finished substrate structure of the first embodiment does not have the first bottom silicon 302a in the first substrate 301a, so the first bottom silicon 302a needs to be removed.

[0282] Since the first silicon substrate 302a is provided with a first silicon oxide layer 303a, when dry etching or grinding reaches the first silicon oxide layer 303a, it means that the first silicon substrate 302a has been completely removed.

[0283] Additionally, refer to Figure 9 Step S42: Etching the first substrate 301a may include, but is not limited to, the following steps:

[0284] Step S421: Etch the first silicon oxide layer 303a and the first top silicon 304a in the first substrate 301a to form a recessed region at the bottom of the first substrate 301a to define the range of the first imaging window 106a.

[0285] In some examples, the first substrate 301a forms the first support 101a of the mesh structure of the first embodiment, and the hollow portion of the first support 101a defines the first imaging window 106a of the mesh structure. Therefore, a recessed region needs to be formed at the bottom of the first substrate 301a by etching the first silicon oxide layer 303a and the first top silicon 304a, and the size of the recessed region is the size of the first imaging window 106a.

[0286] Furthermore, since the first support beam 202 of the carrier structure in the first embodiment is located within the range of the first imaging window 106a, and is formed by etching the first top silicon 304a of the first substrate 301a in a subsequent step, the etching in this step needs to retain the thickness of the first support beam 202 in the first top silicon 304a of the first substrate 301a, so as to facilitate the implementation of subsequent steps.

[0287] Additionally, step S43: fabricating the first support unit 201a may include, but is not limited to, the following steps:

[0288] Step S431: Within the range of the first imaging window 106a, a plurality of first support units 201a for supporting the first support film 102a are prepared using the first top silicon 304a and the first sacrificial layer 305a in the first substrate 301a.

[0289] In some examples, the removal of the first bottom silicon 302a can be performed using a thinning process, the top silicon in the first substrate 301a can be etched using deep silicon etching, and the first sacrificial layer 305a can be etched using VHF or BOE. When there are different levels of first support units 201a, photolithography, development, and photoresist are performed first, followed by ICP or DRIE etching to form the first support units 201a.

[0290] The first support unit 201a includes a first support beam 202 and a second support beam 203 stacked together, with the second support beam 203 positioned between the first support beam 202 and the first support membrane 102a. Therefore, the first support beam 202 is first prepared from the bottom of the carrier structure, and the second support beam 203 is prepared after the first support beam 202 is completed.

[0291] Additionally, refer to Figure 10 Step S43: Fabricating the first support unit 201a may include, but is not limited to, the following steps:

[0292] Step S432: Etch the first top silicon 304a to the first sacrificial layer 305a in the first substrate 301a according to the grid shape to form a plurality of first support beams 202.

[0293] In some examples, since the above steps have already made the first top silicon 304a portion within the first imaging window 106a meet the thickness of the first support beam 202, this step only needs to etch at a preset position according to the mesh shape to meet the requirements of the first support beam 202.

[0294] It is understandable that, since the first sacrificial layer 305a is located on top of the first top silicon 304a, when etching reaches the first sacrificial layer 305a at the preset position, it means that the etching position has penetrated the first top silicon 304a, and the etching process at the current position needs to be stopped.

[0295] Meanwhile, after the etching process of the first top silicon 304a is completed, the shape and position of each first support beam 202 have been determined, and the shape of the hollow part has been determined. The preparation process of the second support beam 203 needs to be carried out in accordance with the first support beam 202.

[0296] Additionally, refer to Figure 11Step S43: Fabricating the first support unit 201a may include, but is not limited to, the following steps:

[0297] Step S433: Etch the first sacrificial layer 305a to the first support film 102a according to the grid shape to form a plurality of second support beams 203, each second support beam 203 and each first support beam 202 corresponding to one another, and stacked to form each first support unit 201a.

[0298] In some examples, the second support beam 203 is formed by etching the first sacrificial layer 305a. In order to ensure that the shape and position of each second support beam 203 corresponds one-to-one with the shape and position of each first support beam 202, the etching process of the first sacrificial layer 305a needs to be carried out within the hollowed-out area defined by the first support beam 202.

[0299] It is understandable that the first support film 102a is located on top of the first sacrificial layer 305a. When the etching of the first sacrificial layer 305a reaches the first support film 102a, the etching process needs to be stopped.

[0300] Specifically, the first sacrificial layer 305a is etched using BOE, or buffered silicon oxide etchant, a mixed acid solution commonly used to remove silicon oxide, typically a mixture of hydrofluoric acid and ammonium fluoride. Using BOE to remove the first sacrificial layer 305a can, to some extent, avoid over-etching other semiconductor materials, and the etching rate is moderate and stable.

[0301] Additionally, refer to Figure 12 Step S44: Remove the first support piece 306, which may include, but is not limited to, the following steps:

[0302] Step S441: Use laser debonding process to remove the first support sheet 306 on top of the first support film 102a and remove the temporary bonded adhesive layer 308.

[0303] In some examples, after the second support beam 203 is formed, the first support body 101a at the bottom of the first support membrane 102a has been prepared. It is necessary to remove the first support sheet 306 at the top of the first support membrane 102a by laser debonding process, and finally remove the temporary bonding adhesive layer 308 to complete the preparation process of the carrier structure of the first embodiment.

[0304] Additionally, refer to Figures 13 to 21 In the preparation process of the second embodiment of the grid structure for cryo-electron microscopy, the step of preparing a support at the bottom of the support film may include, but is not limited to, the following steps:

[0305] Step S45: Remove the second bottom silicon 302b from the second substrate 301b and remove the second sacrificial layer 305b;

[0306] Step S46: Prepare the first metal layer 104;

[0307] Step S47: Prepare a titanium ring having a second metal layer 105 and a second support sheet 401;

[0308] Step S48: Bond the first metal layer 104 and the second metal layer 105;

[0309] Step S49: Remove the first support piece 306 and the second support piece 401.

[0310] In some examples, an etching process is used to remove the second bottom silicon 302b and the second sacrificial layer 305b in the second substrate 301b, thereby exposing the bottom surface of the support film.

[0311] Furthermore, after removing the second bottom silicon 302b and the second sacrificial layer 305b, a first metal layer 104 can be prepared on the bottom surface of the support film using processes such as deposition. The first metal layer 104 is used to connect the titanium ring prepared in subsequent processes.

[0312] Simultaneously, during the preparation of the titanium ring, a bonding process is used to connect the titanium sheet and the second support sheet 401, and a second metal layer 105 is deposited on the titanium sheet. After depositing the second metal layer 105, the shape of the second imaging window 106b is cut out on the titanium sheet using cutting processes such as laser cutting, thereby completing the preparation of the titanium ring.

[0313] Furthermore, the first metal layer 104 and the second metal layer 105 are bonded using a bonding process, so that the titanium ring and the support film are connected as one unit. Both the titanium ring and the metal layers have good conductivity, thereby improving the conductivity of the carrier structure.

[0314] Understandably, the first support sheet 306 and the second support sheet 401 need to be removed using processes such as laser debonding to complete the fabrication of the carrier structure.

[0315] Additionally, refer to Figure 15 Step S2: After etching the support film to form at least one accommodating channel, the following steps may be included but not limited to:

[0316] Step S23: Prepare a support structure on top of the second support membrane 102b according to the grid shape to form a plurality of second support units 201b for supporting the second support membrane 102b.

[0317] In some examples, the support structure of the second embodiment is located on top of the second support film 102b and is independent of the second substrate 301b at the bottom of the second support film 102b. The second substrate 301b at the bottom of the second support film 102b can provide support for the second support film 102b, which facilitates the fabrication of the support structure on top of the second support film 102b.

[0318] It is understandable that when the support structure is prepared, each of the second accommodating channels 1021b on the second support membrane 102b has been prepared. The second support unit 201b in the support structure needs to be prepared according to the grid shape and the position of each of the second accommodating channels 1021b to avoid the second support unit 201b covering the top of the second accommodating channel 1021b.

[0319] Additionally, step S23: preparing a support structure on top of the second support membrane 102b according to the mesh shape may include, but is not limited to, the following steps:

[0320] Step S231: The support structure is prepared using a lift-off process.

[0321] In some examples, the lift-off process is a widely used patterning technique in the field of micro- and nanofabrication, primarily used to fabricate precise patterns of metals or other materials on a substrate. The principle involves using photoresist or other second sacrificial layer 305b material as a mask to deposit a thin film onto the substrate and the second sacrificial layer 305b. The second sacrificial layer 305b and the thin film material on it are then removed, leaving the thin film material in the areas of the substrate not covered by the second sacrificial layer 305b, thus forming the desired pattern.

[0322] Additionally, refer to Figure 16 In some examples, when fabricating the second support 101b at the bottom of the second support film 102b, it is necessary to support the second support film 102b on top of it to prevent the state of the second support film 102b from being affected. Therefore, the first support sheet 306 is bonded to the top of the second support film 102b and the support structure using a temporary bonding process. It is understood that a laser debonding layer 307 and a temporary bonding adhesive layer 308 also exist between the first support sheet 306 and the support structure.

[0323] Additionally, step S45: removing the second bottom silicon 302b from the second substrate 301b and removing the second sacrificial layer 305b may include, but is not limited to, the following steps:

[0324] Step S451: Remove the second bottom silicon 302b of the second substrate 301b by dry etching or polishing;

[0325] Step S452: Remove the second sacrificial layer 305b by wet etching.

[0326] In some examples, the second sacrificial layer 305b and the second bottom silicon 302b in the second substrate 301b are not present in the finished net structure of the second embodiment and need to be removed. Here, dry etching and wet etching are used to remove the second bottom silicon 302b and the second sacrificial layer 305b, respectively.

[0327] Additionally, refer to Figure 17 Step S46: Prepare the first metal layer 104, which may include, but is not limited to, the following steps:

[0328] Step S461: Prepare a first metal layer 104 at the bottom of the second support film 102b.

[0329] In some examples, since the second support 101b of the carrier structure in the second embodiment is made of titanium, it is difficult for the titanium to be directly bonded to the second support film 102b. Therefore, the first metal layer 104 is needed to assist the titanium material and the second support film 102b in bonding.

[0330] In this embodiment, the first metal layer 104 is retained in the finished mesh structure and is located within the second support 101b, which defines the second imaging window 106b of the mesh structure. Therefore, the first metal layer 104 is formed at the bottom edge of the second support film 102b and surrounds the second imaging window 106b.

[0331] In some examples, the titanium sheet is first processed into a shape that conforms to the second imaging window 106b before the titanium and the second support film 102b are bonded together. Since the first metal layer 104 defining the second imaging window 106b is approximately annular, the titanium sheet is also processed into a titanium ring.

[0332] Additionally, refer to Figure 18 Step S47: Preparing a titanium ring having a second metal layer 105 and a second support sheet 401 may include, but is not limited to, the following steps:

[0333] Step S471: Use a temporary bonding process to bond the titanium sheet to the second support sheet 401;

[0334] Step S472: Deposit a second metal layer 105 on the titanium sheet.

[0335] In some examples, the titanium sheet needs to maintain its shape during processing, so a second support sheet 401 is attached to the titanium sheet to provide sufficient support.

[0336] Furthermore, since titanium is difficult to bond directly with the first metal layer 104, a second metal layer 105 is deposited on top of the titanium sheet. The second metal layer 105 can be easily bonded with the first metal layer 104, so the titanium sheet and the second support film 102b are bonded through the first metal layer 104 and the second metal layer 105.

[0337] Additionally, refer to Figure 19 Step S47: Preparing a titanium ring having a second metal layer 105 and a second support sheet 401 may include, but is not limited to, the following steps:

[0338] Step S473: Cut the second metal layer 105 and the titanium sheet on the second support sheet 401 to form a ring structure corresponding to the shape of the first metal layer 104.

[0339] In some examples, the shape of the second imaging window 106b is cut out on the second metal layer 105 and the titanium sheet. Since the shape of the second imaging window 106b has been defined by the first metal layer 104, the titanium sheet and the second metal layer 105 need to be cut according to the shape of the first metal sheet to form a titanium ring.

[0340] Additionally, step S473: cutting the second metal layer 105 and the titanium sheet on the second support sheet 401 may include, but is not limited to, the following steps:

[0341] Step S4731: Use laser cutting to cut the second metal layer 105 and the titanium sheet.

[0342] Additionally, refer to Figure 20 In some examples, the first metal layer 104 and the second metal layer 105 can be made of metal materials such as Al / Ge, Au / Sn, Au, Cu, Al, etc., to facilitate the bonding process of the first metal layer 104 and the second metal layer 105.

[0343] Furthermore, the first metal layer 104 and the second metal layer 105 do not need to be separated after bonding. Therefore, the first metal layer 104 and the second metal layer 105 can be permanently bonded or temporarily bonded.

[0344] Additionally, step S48: bonding the first metal layer 104 and the second metal layer 105 may include, but is not limited to, the following steps:

[0345] Step S481: The first metal layer 104 and the second metal layer 105 are bonded by eutectic bonding or metal hot pressing bonding.

[0346] In some examples, eutectic bonding is a bonding method based on the direct transformation of a eutectic alloy from a solid to a liquid state under specific temperature and pressure conditions, followed by solidification of the liquid alloy at the interface, thereby achieving a strong bond between two surfaces to be bonded.

[0347] Furthermore, hot-press bonding of metals is primarily based on the plastic deformation and diffusion mechanisms of metals. Permanent bonding is achieved under the combined action of heating and pressure.

[0348] Additionally, refer to Figure 21 Step S49: Removing the first support piece 306 and the second support piece 401 may include, but is not limited to, the following steps:

[0349] Step S491: Remove the first support sheet 306 and the second support sheet 401 using a laser debonding process.

[0350] In some examples, the second support 101b of the mesh structure in the second embodiment is completed after the titanium ring is bonded to the second support film 102b. The fabrication process of the mesh structure can be completed by removing the first support sheet 306 and the second support sheet 401 using a laser debonding process.

[0351] In addition, in the preparation process of the third embodiment of the carrier structure for cryo-electron microscopy, step S1: preparing a substrate and preparing a support film on top of the substrate includes, but is not limited to, the following steps:

[0352] Reference Figure 25 Step S14: Create conductive material filling area 402;

[0353] Reference Figure 26 Step S15: Deposit metal into the conductive material filling region 402.

[0354] In some examples, an etching process is used to etch downwards from the third support film 102c to form the conductive fill region 402. After the conductive fill region 402 is prepared, a deposition process is used to deposit metal onto the conductive fill region 402 to ensure the conductivity of the carrier structure.

[0355] Additionally, step S14: the fabrication of the conductive material filling region 402 includes, but is not limited to, the following steps:

[0356] Step S141: Etch the third support film 102c and the third substrate 301c down to the bottom of the second silicon oxide layer 303b of the third substrate 301c to form the conductive filling region.

[0357] In some examples, the conductive material filling region 402 is located inside the second top silicon layer 304b and the second silicon oxide layer 303b in the third substrate 301c. To facilitate the deposition of metal material in the conductive material filling region 402, both the third support film 102c and the third substrate 301c are provided with corresponding holes. The holes on the third support film 102c and the third substrate 301c correspond one-to-one, thereby forming a deposition channel. It can be understood that the end region of the deposition channel is the conductive material filling region 402.

[0358] Furthermore, corresponding holes are formed on the third support film 102c and the third substrate 301c by etching, and the etching stops at the junction of the second silicon oxide layer 303b and the third bottom silicon 302c in the third substrate 301c, ensuring that the conductive material filling area 402 extends to the top surface of the third bottom silicon 302c.

[0359] Additionally, step S15: depositing metal in the conductive fill region 402 includes, but is not limited to, the following steps:

[0360] Step 151: Deposit a conductor 403 in the conductive filling area, wherein the conductor 403 is one of aluminum, titanium nitride, tungsten, or gold.

[0361] In some examples, metallic materials such as aluminum, titanium nitride, tungsten, and gold are conductive and suitable for the fabrication process of the network structure. When conductors such as aluminum, titanium nitride, tungsten, and gold are deposited in the conductive filling region 402, the resistivity of the network structure can be reduced.

[0362] Meanwhile, the absence of a sacrificial layer in the grid structure of the third embodiment also reduces the resistivity of the grid structure. Therefore, the preparation method without a sacrificial layer combined with the deposition method of conductive materials can improve the conductivity of the grid structure, enhance imaging quality, and provide strong support for high-precision cryo-electron microscopy observations.

[0363] Additionally, step S151: after depositing the conductive material 403 in the conductive material filling region, includes, but is not limited to, the following steps:

[0364] Step S1511: Perform annealing to form an ohmic contact between the conductor 403 and the third substrate 301c.

[0365] In some examples, the conductor 403 needs to form an electrical connection with the structure in the third substrate 301c. However, the structure in the third substrate 301c is mostly semiconductor. Therefore, it is necessary to form ohmic contacts between the conductor 403 and the structure in the third substrate 301c by rapid annealing.

[0366] Furthermore, rapid annealing can induce a reaction between the metal and semiconductor surfaces, forming low-resistivity metal silicides. These silicides have low resistivity, which can effectively reduce the contact resistance between the metal and semiconductor, thereby making the metal-semiconductor contact closer to the characteristics of an ohmic contact.

[0367] In some examples, refer to Figure 27 After the conductor 403 is deposited, the third accommodating channel 1021c can be fabricated on the surface of the third support film 102c by etching or other methods. (Refer to...) Figure 28 After the third accommodating channel 1021c is prepared, a first support sheet 306 is set on the top of the carrier structure. The first support sheet 306 can be set by a temporary bonding process.

[0368] Additionally, step S4: preparing a support at the bottom of the support membrane includes, but is not limited to, the following steps:

[0369] Reference Figure 29 Step S50: Deposit a silicon oxide film on the bottom of the third substrate 301c;

[0370] Reference Figure 30 Step S51: Etch the third bottom silicon 302c and the second silicon oxide layer 303b of the third substrate 301c to form the third support beam 204;

[0371] Reference Figure 31 Step S52: Remove the first support piece 306;

[0372] Reference Figure 32 Step S53: Etch the second top silicon 304b of the third substrate 301c to form the fourth support beam 205.

[0373] In some examples, a silicon oxide film needs to be deposited on the bottom of the third substrate 301c. After the silicon oxide film is deposited, the third bottom silicon 302c and the second silicon oxide layer 303b in the third substrate 301c are etched to form the third support beam 204. Further, the first support sheet 306 at the top of the carrier structure is removed, and finally the fourth support beam 205 is formed by etching the second top silicon 304b in the third substrate 301c.

[0374] The third support beam 204 enhances the overall strength of the mesh structure, facilitating a reduction in the thickness of the fourth support beam 205. Without the third support beam 204, the thickness of the fourth support beam 205 would need to be between 15 μm and 50 μm. In the third embodiment of this application, the inclusion of the third support beam 204 allows for a reduction in the thickness of the fourth support beam 205 to between 2 μm and 5 μm. This reduction in the thickness of the fourth support beam 205 helps eliminate the obstruction of the observation area by shadows during electron microscopy, significantly improving the clarity and integrity of the observation field.

[0375] Additionally, step S50 involves depositing a silicon oxide film on the bottom of the third substrate 301c, including but not limited to the following steps:

[0376] Step S501: Deposit silicon oxide film using ICP-VD process.

[0377] In some examples, ICPCVD can deposit thin films at lower temperatures compared to conventional thermochemical vapor deposition. This is because plasma provides the additional energy to promote chemical reactions, allowing them to proceed rapidly at lower temperatures, thus reducing thermal damage to the substrate material. Simultaneously, the deposited films exhibit higher density and uniformity with fewer defects due to ion bombardment. Ions can also sputter-clean the film surface, removing contaminants and impurities, further improving film quality. Understandably, by precisely controlling plasma parameters, precise control over film thickness, composition, structure, and other properties can be achieved.

[0378] Additionally, step S52: removing the first support piece 306 includes, but is not limited to, the following steps:

[0379] Step S521: Remove the first support sheet 306 by acrylic acid cleaning or IPA cleaning.

[0380] When the first support sheet 306 is temporarily bonded to the top of the carrier structure, it can be removed by a debonding process, which includes acrylic cleaning or IPA cleaning. Alternatively, the first support sheet 306 can also be bonded to the top of the carrier structure using UV adhesive. UV adhesive bonding utilizes ultraviolet light to cure the adhesive, thereby connecting the carrier structure and the first support sheet 306.

[0381] It is worth noting that etching processes are used multiple times in each step of this application to remove excess material, and photolithography can be used before each etching step to ensure the accuracy of the etching process. Specifically, during the photolithography process, photoresist can form a precise pattern template on the carrier structure, and the subsequent etching process will be carried out according to this template, thereby achieving precise control of the carrier structure.

[0382] The embodiments of this application have been described in detail above with reference to the accompanying drawings. However, this application is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of this application. Furthermore, unless otherwise specified, the embodiments and features described in the embodiments of this application can be combined with each other.

Claims

1. A method for preparing a carrier network structure, characterized in that, include: A substrate (301) is prepared, and a support film is prepared on top of the substrate (301); The support film is etched to form at least one accommodating channel (1021); A first support sheet (306) is provided on top of the support membrane; A support is prepared at the bottom of the support membrane.

2. The method for preparing the carrier network structure according to claim 1, characterized in that, The preparation of the substrate (301) and the preparation of a support film on top of the substrate (301) includes: A sacrificial layer (305) is prepared on the surface of the substrate (301); A support film is prepared on the surface of the sacrificial layer (305).

3. The method for preparing the carrier network structure according to claim 1, characterized in that, The substrate preparation (301) includes: The substrate (301) is prepared using silicon material.

4. The method for preparing the carrier network structure according to claim 2, characterized in that, The preparation of the sacrificial layer (305) on the surface of the substrate (301) includes: The sacrificial layer (305) is prepared using phosphosilicate glass, borosilicate glass, or silicon oxide.

5. The method for preparing the carrier network structure according to claim 1, characterized in that, Etching the support film to form at least one accommodating channel (1021) includes: Frontal exposure microscopy; The support film is etched using a dry etching process to form the accommodating channel (1021).

6. The method for preparing the carrier network structure according to claim 1, characterized in that, A first support sheet (306) is disposed on top of the support membrane, comprising: The first support sheet (306) is prepared using a light-transmitting material.

7. The method for preparing the carrier network structure according to claim 1 or 5, characterized in that, A first support sheet (306) is disposed on top of the support membrane, comprising: The first support sheet (306) is prepared using a material that can withstand at least 400°C.

8. The method for preparing the carrier network structure according to claim 2, characterized in that, A support is prepared at the bottom of the support membrane, comprising: Remove the first bottom silicon (302a) from the first substrate (301a); Etch the first substrate (301a); Fabrication of the first support unit (201a); Remove the first support piece (306).

9. The method for preparing the carrier network structure according to claim 8, characterized in that, A first support sheet (306) is disposed on top of the support membrane, comprising: A temporary bonding adhesion layer (308) and a laser debonding layer (307) are formed on top of the first support film; The first support sheet (306) is bonded to the top of the laser debonding layer (307).

10. The method for preparing the carrier network structure according to claim 8, characterized in that, Removing the first bottom silicon (302a) from the first substrate (301a) includes: The first bottom silicon (302a) in the first substrate (301a) is removed by dry etching or polishing.

11. The method for preparing the carrier network structure according to claim 8, characterized in that, Etching the first substrate (301a) includes: The first silicon oxide layer (303a) and the first top silicon (304a) in the first substrate (301a) are etched to form a recessed region at the bottom of the first substrate (301a) to define the range of the first imaging window (106a).

12. The method for preparing the carrier network structure according to claim 11, characterized in that, The fabrication of the first support unit (201a) includes: Within the first imaging window (106a), a plurality of first support units (201a) for supporting the first support film are prepared using the first top silicon (304a) and the first sacrificial layer (305a) in the first substrate (301a).

13. The method for preparing the carrier network structure according to claim 12, characterized in that, The fabrication of the first support unit (201a) includes: According to the grid shape, the first top silicon (304a) in the first substrate (301a) to the first sacrificial layer (305a) are etched to form a plurality of first support beams (202).

14. The method for preparing the carrier network structure according to claim 13, characterized in that, The fabrication of the first support unit (201a) includes: The first sacrificial layer (305a) is etched to the first support film according to the grid shape to form a plurality of second support beams (203), each second support beam (203) and each first support beam (202) correspond one-to-one, and are stacked to form each first support unit (201a).

15. The method for preparing the carrier network structure according to claim 9, characterized in that, The removal of the first support sheet (306) includes: The first support sheet (306) on top of the first support film is removed by laser debonding process, and the temporary bonded adhesive layer (308) is also removed.

16. The method for preparing the carrier network structure according to claim 2, characterized in that, A support is prepared at the bottom of the support membrane, comprising: Remove the second bottom silicon (302b) from the second substrate (301b) and remove the second sacrificial layer (305b); Prepare the first metal layer (104); A titanium ring having a second metal layer (105) and a second support sheet (401) is prepared; Bond the first metal layer (104) and the second metal layer (105); Remove the first support piece (306) and the second support piece (401).

17. The method for preparing the carrier network structure according to claim 16, characterized in that, After etching the support film to form at least one accommodating channel (1021), the method further includes: A support structure is prepared on top of the second support membrane (102b) according to the grid shape to form a plurality of second support units (201b) for supporting the second support membrane (102b).

18. The method for preparing the carrier network structure according to claim 17, characterized in that, The preparation of the support structure on top of the second support membrane (102b) according to the mesh shape includes: The support structure was fabricated using a lift-off process.

19. The method for preparing the carrier network structure according to claim 16, characterized in that, Removing the second bottom silicon (302b) from the second substrate (301b) and removing the second sacrificial layer (305b) includes: The second bottom silicon (302b) of the second substrate (301b) is removed by dry etching or polishing; The second sacrificial layer (305b) was removed by wet etching.

20. The method for preparing the carrier network structure according to claim 16, characterized in that, The preparation of the first metal layer (104) includes: A first metal layer (104) is prepared at the bottom of the second support film (102b).

21. The method for preparing the carrier network structure according to claim 16, characterized in that, The preparation of the titanium ring having a second metal layer (105) and a second support sheet (401) includes: A temporary bonding process is used to bond the titanium sheet to the second support sheet (401); A second metal layer (105) is deposited on the titanium sheet.

22. The method for preparing the carrier network structure according to claim 21, characterized in that, The preparation of the titanium ring having a second metal layer (105) and a second support sheet (401) includes: Cut the second metal layer (105) and the titanium sheet on the second support sheet (401) to form a ring structure corresponding to the shape of the first metal layer (104).

23. The method for preparing the carrier network structure according to claim 22, characterized in that, Cutting the second metal layer (105) and the titanium sheet on the second support sheet (401) includes: The second metal layer (105) and the titanium sheet are cut using a laser.

24. The method for preparing the carrier network structure according to claim 16, characterized in that, Bonding the first metal layer (104) and the second metal layer (105) includes: The first metal layer (104) and the second metal layer (105) are bonded by eutectic bonding or metal hot pressing bonding.

25. The method for preparing the carrier network structure according to claim 22, characterized in that, Removing the first support sheet (306) and the second support sheet (401) includes: The first support sheet (306) and the second support sheet (401) are removed by laser debonding process.

26. The method for preparing the carrier network structure according to claim 1, characterized in that, The preparation of the substrate (301) and the preparation of a support film on top of the substrate (301) includes: Fabricate the conductive material filling region (402); Metal is deposited in the conductive material filling region (402).

27. The method for preparing the carrier network structure according to claim 26, characterized in that, The fabrication of the conductive material filling region (402) includes: The third support film (102c) and the third substrate (301c) are etched down to the bottom of the silicon dioxide layer of the third substrate (301c) to form the conductive material filling region (402).

28. The method for preparing the carrier network structure according to claim 26, characterized in that, The step of depositing metal in the conductive fill region (402) includes: A conductor (403) is deposited in the conductive filling region (402), wherein the conductor (403) is one of aluminum, titanium nitride, tungsten, or gold.

29. The method for preparing the carrier network structure according to claim 28, characterized in that, After depositing the conductor (403) in the conductive filling region (402), the process includes: Annealing is performed to form an ohmic contact between the conductor (403) and the third substrate (301c).

30. The method for preparing the carrier network structure according to claim 26, characterized in that, A support is prepared at the bottom of the support membrane, comprising: A silicon oxide film is deposited on the bottom of the third substrate (301c); The third bottom silicon (302c) and the second silicon oxide layer (303b) of the third substrate (301c) are etched to form the third support beam (204); Remove the first support piece (306); The second top silicon (304b) of the third substrate (301c) is etched to form the fourth support beam (205).

31. The method for preparing the carrier network structure according to claim 30, characterized in that, A silicon oxide film is deposited on the bottom of the third substrate (301c), including: Silicon oxide film was deposited using the ICP-VD process.

32. The method for preparing the carrier network structure according to claim 30, characterized in that, Removing the first support sheet (306) includes: The first support sheet (306) is removed by acrylic cleaning or IPA cleaning.

33. A grid structure for cryo-electron microscopy, characterized in that, include: Support structure; A support membrane having at least one through-hole receiving channel (1021), the support membrane being made of silicon nitride, silicon carbide or nickel-titanium alloy; An interlayer is located between the support film and the support body, the hollow portion of the interlayer and the support body defines an imaging window, and the accommodating channel (1021) is located within the area of ​​the imaging window.

34. The grid structure for cryo-electron microscopy according to claim 33, characterized in that, The thickness of the support film is greater than or equal to 15 nm and less than or equal to 50 nm.

35. The grid structure for cryo-electron microscopy according to claim 33, characterized in that, The thickness of the support is greater than or equal to 20 μm and less than or equal to 100 μm.

36. The grid structure for cryo-electron microscopy according to claim 33, characterized in that, The grid structure for cryo-electron microscopy further includes a support structure, which is attached to the support membrane and is used to support the support membrane.

37. The grid structure for cryo-electron microscopy according to claim 36, characterized in that, The support structure has several hollowed-out sections, and the receiving channel (1021) is located within the range of each hollowed-out section.

38. The grid structure for cryo-electron microscopy according to claim 37, characterized in that, The support structure includes a plurality of support units, which are staggered within the range of the imaging window to form a grid structure, wherein the hollow portion of the grid structure defines each of the cutout portions.

39. The grid structure for cryo-electron microscopy according to claim 38, characterized in that, The thickness of the support structure is greater than or equal to 1 μm and less than or equal to 5 μm.

40. The grid structure for cryo-electron microscopy according to claim 39, characterized in that, The interlayer is made of phosphosilicate glass, borosilicate glass, or silicon oxide.

41. The grid structure for cryo-electron microscopy according to claim 40, characterized in that, The thickness of the interlayer is greater than or equal to 100 nm and less than or equal to 800 nm.

42. The grid structure for cryo-electron microscopy according to claim 40, characterized in that, The support unit includes a second support beam (203), each of the second support beams (203) is attached to the bottom of the support membrane, and adjacent second support beams (203) form a second grid.

43. The grid structure for cryo-electron microscopy according to claim 42, characterized in that, Each of the second support beams (203) is formed by etching the interlayer.

44. The grid structure for cryo-electron microscopy according to claim 42, characterized in that, The support is made of silicon.

45. The grid structure for cryo-electron microscopy according to claim 44, characterized in that, The support unit further includes a first support beam (202), each of the first support beams (202) is attached to the bottom of each of the second support beams (203), and adjacent first support beams (202) form a first grid, each of the first grids and each of the second grids are connected in a one-to-one correspondence to form each of the hollow parts.

46. ​​The grid structure for cryo-electron microscopy according to claim 45, characterized in that, Each of the first support beams (202) is formed by etching the support body.

47. The grid structure for cryo-electron microscopy according to claim 46, characterized in that, The thickness of the first support beam (202) is less than the thickness of the support body.

48. The grid structure for cryo-electron microscopy according to claim 39, characterized in that, The interlayer is made of metal.

49. The grid structure for cryo-electron microscopy according to claim 48, characterized in that, The interlayer includes a first metal layer (104) disposed at the bottom of the support membrane and a second metal layer (105) disposed at the top of the support, wherein the first metal layer (104) and the second metal layer (105) are bonded together.

50. The grid structure for cryo-electron microscopy according to claim 49, characterized in that, The support is made of titanium.

51. The grid structure for cryo-electron microscopy according to claim 48, characterized in that, The support structure is made of titanium and is attached to the top of the support membrane.

52. The grid structure for cryo-electron microscopy according to claim 51, characterized in that, The width of the support unit is greater than or equal to 10 μm and less than or equal to 20 μm.

53. The grid structure for cryo-electron microscopy according to claim 39, characterized in that, The interlayer includes a top silicon layer and a silicon oxide layer stacked together, the silicon oxide layer being located between the top silicon layer and the support, and the interlayer having a deposition channel for depositing a conductor (403).

54. The grid structure for cryo-electron microscopy according to claim 53, characterized in that, The conductor (403) is made of one of the following materials: aluminum, titanium nitride, tungsten, or gold.

55. The grid structure for cryo-electron microscopy according to claim 53, characterized in that, The support unit includes a fourth support beam (205), each of the fourth support beams (205) is attached to the bottom of the support membrane, and adjacent fourth support beams (205) form a fourth grid.

56. The grid structure for cryo-electron microscopy according to claim 55, characterized in that, The fourth grid is formed by etching the top silicon.

57. The grid structure for cryo-electron microscopy according to claim 55, characterized in that, The support unit also includes a third support beam (204), which is attached to the bottom of the silicon oxide layer. Adjacent third support beams (204) form a third grid, and each third grid and each fourth grid are connected in a one-to-one correspondence to form a hollow part.

58. The grid structure for cryo-electron microscopy according to claim 57, characterized in that, The third grid is formed by etching the support.

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