Sample preparation device, sample preparation system and sample preparation method

Abstract

The application relates to a sample preparation device, a sample preparation system and a sample preparation method. The sample preparation device comprises a cooling mechanism, a low-temperature heat sink and a heat conduction mechanism. The low-temperature heat sink is used for carrying a sample carrying unit. The heat conduction mechanism is connected between the cooling mechanism and the low-temperature heat sink. The cooling mechanism cools the sample carrying unit carried on the low-temperature heat sink through the heat conduction mechanism and the low-temperature heat sink. The heat conduction mechanism has a good heat conduction effect. The cold energy of the cooling mechanism is quickly conducted to the low-temperature heat sink through the heat conduction mechanism. The temperature of the low-temperature heat sink can be accurately controlled. The temperature of the sample carrying unit can be accurately controlled. The sample preparation efficiency is high. The arrangement of the cooling mechanism and the low-temperature heat sink is flexible. The reliability of the device is increased.

Description

Technical Field

[0001] This application relates to the field of cryo-electron microscopy, and in particular to a sample preparation apparatus, a sample preparation system, and a sample preparation method. Background Technology

[0002] Transmission electron microscopy (TEM) is widely used in materials science, physics, and biochemistry. During TEM observation, a sample support chip is used to hold the sample. The sample support chip typically includes a support film, which is an amorphous thin film with a thickness of, for example, 20 mm to 40 mm. In cryo-electron microscopy techniques, the support film is often porous, and the solution carrying the sample forms a film at the pores, allowing the sample to remain within them. The electron beam emitted by the TEM then passes through the sample, thus creating an image.

[0003] Samples in related technologies typically include the following sample preparation methods:

[0004] Method 1 involves overloading a sample liquid onto a sample carrier chip during sample preparation. The liquid is then thinned using methods such as natural evaporation, plasma cooling, or electrospray printing. However, existing sample preparation devices are merely theoretical concepts and have not been practically implemented, resulting in low reliability. Furthermore, while the device uses airflow to cool the sample liquid during preparation, it cannot precisely control the sample thickness on the carrier chip. Additionally, these devices cannot rapidly reduce sample temperature, and temperature control accuracy is low. The process is time-consuming and inefficient.

[0005] Method two involves using a pump or gas to drive the fluid sample through the sample chamber of the sample-carrying chip, filling the entire chamber. The device then cools the liquid within the chamber. However, the fluid sample cannot actually completely fill the chamber, making effective control of sample thickness impossible. Furthermore, the devices in this method cannot rapidly reduce sample temperature, resulting in low temperature control accuracy. Sample preparation is also time-consuming and inefficient. Summary of the Invention

[0006] Based on this, this application aims to solve at least one of the technical problems existing in the prior art, and to provide a sample preparation device, a sample preparation system and a sample preparation method.

[0007] In a first aspect, this application provides a sample preparation apparatus, comprising:

[0008] Cooling mechanism;

[0009] A low-temperature heat sink is used to support the sample carrier unit.

[0010] A heat conduction mechanism is provided, which is connected between the cooling mechanism and the low-temperature heat sink. The cooling mechanism cools the sample carrier unit supported on the low-temperature heat sink through the heat conduction mechanism and the low-temperature heat sink.

[0011] In one embodiment, the heat conduction mechanism includes a cooling braided strip connected between the cooling mechanism and the low-temperature heat sink.

[0012] In one embodiment, the cooling braided tape is made of at least one material including oxygen-free copper or graphite sheets.

[0013] In one embodiment, the cooling mechanism includes a liquid gas container.

[0014] In one embodiment, the liquid gas container is provided with a liquid storage cavity, the top of which is open so that the liquid gas inside the liquid storage cavity is exposed through the top of the liquid storage cavity.

[0015] In one embodiment, the sample preparation device further includes a support frame arranged adjacent to the liquid gas container, and the cryogenic heat sink is installed on top of the support frame.

[0016] In one embodiment, the support frame includes legs and a heat insulation base, the legs being connected to the heat insulation base, and the low-temperature heat sink being mounted on the heat insulation base.

[0017] In one embodiment, the sample preparation device further includes a base, and the cooling mechanism is disposed on the base.

[0018] Secondly, this application provides a sample preparation system, including the aforementioned sample preparation device, and further including a sample carrying unit; the sample carrying unit includes:

[0019] A heating mechanism for supporting a sample carrier chip and heating the sample carrier chip;

[0020] A support platform, the support platform being used to support the heating mechanism; and

[0021] A sample carrier chip includes a substrate and a support member. The support member is connected to the substrate and has a chamber and a sample inlet. The inner wall of the chamber is hydrophilic, and the sample inlet is connected to the chamber.

[0022] In one embodiment, the heating mechanism is provided with a positioning groove for positioning the sample chip, and the sample chip is disposed in the positioning groove and bonded and fixed to the heating mechanism.

[0023] In one embodiment, at least a portion of the support platform is disposed on the low-temperature heat sink, and another portion of the support platform extends out of the low-temperature heat sink and is suspended in the air. The projection of the sample chip along the thickness direction of the support platform is offset from that of the low-temperature heat sink.

[0024] In one embodiment, the sample preparation device further includes a positioning frame, a portion of which is connected to the low-temperature heat sink, and another portion of which extends out of the low-temperature heat sink and is suspended in the air. The positioning frame is used to support and position the support platform.

[0025] In one embodiment, the sample carrier unit further includes a clamping member connected to the carrier stage, the clamping member being used to press and fix the sample chip and the heating mechanism to the carrier stage.

[0026] In one embodiment, the heating mechanism includes a heating chip; the sample carrying unit further includes an adapter plate and an electrical connection connector plate; the heating chip is electrically connected to the adapter plate, and the heating chip is stacked on the adapter plate; the adapter plate is disposed on the carrying platform; the adapter plate is electrically connected to the electrical connection connector plate, and the electrical connection connector plate is used for electrical connection with the controller.

[0027] In one embodiment, the adapter plate is encapsulated and fixed to the carrier platform.

[0028] In one embodiment, the heating chip is electrically connected and thermally contacted to the metal leads of the adapter plate by bonding.

[0029] In one embodiment, the outer wall of the support is provided as a hydrophobic wall surface.

[0030] In one embodiment, the chamber is provided with a plurality of preparation unit areas, and the spacing between the two opposite inner walls of each preparation unit area along the thickness direction of the support is different or the same.

[0031] In one embodiment, the support includes a first support film and a second support film disposed at a distance from each other. The first support film is connected to the substrate, and the second support film is located on the side of the first support film opposite to the substrate and cooperates with the first support film to form the cavity. The first support film is provided with a first observation portion, and the second support film is provided with a second observation portion. The second observation portion is arranged opposite to the first observation portion.

[0032] In one embodiment, the substrate has a first cutout portion, and the projections of the first observation portion and the second observation portion along the thickness direction of the support member are both located within the outline range of the first cutout portion.

[0033] In one embodiment, the first hollow portion has at least two hollow areas, and a reinforcing portion is provided between any two adjacent hollow areas, the reinforcing portion being connected to the inner wall of the first hollow portion.

[0034] In one embodiment, there are multiple first observation units and multiple second observation units; each first observation unit is correspondingly arranged with each of the preparation unit regions, and each second observation unit is correspondingly arranged with each of the preparation unit regions.

[0035] In one embodiment, both the first observation portion and the second observation portion are elongated, and the width of both the first observation portion and the second observation portion is set to 0.6μm-2.5μm.

[0036] In one embodiment, the first observation portion includes a first hole penetrating the first support membrane, and the second observation portion includes a second hole penetrating the second support membrane.

[0037] In one embodiment, the first support membrane includes a first main support portion and a first auxiliary support portion, the first auxiliary support portion being connected to the first main support portion, and the first hole being disposed in the first auxiliary support portion; the thickness of the first auxiliary support portion is less than the thickness of the first main support portion; and / or...

[0038] The second support membrane includes a second main support portion and a second auxiliary support portion, the second auxiliary support portion being connected to the second main support portion, and the second hole being disposed in the second auxiliary support portion; the thickness of the second auxiliary support portion is less than the thickness of the second main support portion.

[0039] In one embodiment, the first main support portion is circumferentially arranged around the first auxiliary support portion; and / or, the second main support portion is circumferentially arranged around the second auxiliary support portion.

[0040] In one embodiment, the first observation unit further includes a first sealing film connected to the first support film, the first sealing film being used to seal the first hole, the first sealing film being transparent under electron beam irradiation; and / or, the second observation unit further includes a second sealing film connected to the second support film, the second sealing film being used to seal the second hole, the second sealing film being transparent under electron beam irradiation.

[0041] In one embodiment, the first sealing film comprises a graphene material; and / or, the second sealing film comprises a graphene material.

[0042] In one embodiment, the support further includes a support column connected between the first support membrane and the second support membrane.

[0043] In one embodiment, the support column is integrally formed with at least one of the first support membrane and the second support membrane.

[0044] In one embodiment, the substrate has a support layer on the side opposite to the support member.

[0045] In one embodiment, the substrate is provided with a sample inlet channel that penetrates the substrate along the thickness direction and is in communication with the sample inlet hole.

[0046] In one embodiment, the substrate is further provided with a flow channel, which is disposed between the sample inlet channel and the sample inlet hole; the flow channel is provided in at least two stages and connected in series; along the sample liquid flow direction, the first flow channel is connected to the sample inlet channel, and the last flow channel is connected to the sample inlet hole.

[0047] In one embodiment, the cross-sectional dimensions of the flow channels at different levels decrease along the direction of sample liquid flow.

[0048] In one embodiment, a recess is formed on the side of the substrate facing the support member, the recess being recessed in a direction away from the support member, and the recess cooperating with the support member to form the flow channel.

[0049] In one embodiment, at least two stages of the flow channels are spirally arranged along the outer periphery of the support, and the distance between the last flow channel and the center of the support is less than the distance between the first flow channel and the center of the support.

[0050] In one embodiment, the sample preparation system further includes an ohmic contact resistor; the ohmic contact resistor is electrically connected to at least one of the substrate and the support; the ohmic contact resistor is configured as two, the two ohmic contact resistors are arranged at intervals, and the two ohmic contact resistors are respectively electrically connected to the positive and negative terminals of an external conductive element.

[0051] Thirdly, this application also provides a sample preparation method for the aforementioned sample preparation system, comprising:

[0052] The heating mechanism is activated to heat the sample carrier chip, and the temperature of the sample carrier chip is controlled to be higher than the first preset temperature.

[0053] The sample liquid is added into the chamber through the inlet port, and the sample liquid spontaneously wets the chamber through capillary action.

[0054] Once the sample liquid wets the entire chamber, the heating mechanism stops working, and the cooling mechanism lowers the temperature of the sample carrier chip to below the second preset temperature, causing the sample liquid inside the sample carrier chip to freeze rapidly.

[0055] The sample preparation device, system, and method described above have a heat conduction mechanism with good heat conduction effect. The cooling capacity of the cooling mechanism is quickly conducted to the low-temperature heat sink through the heat conduction mechanism, which can accurately control the temperature of the low-temperature heat sink and thus accurately control the temperature of the sample carrying unit, ensuring high sample preparation efficiency. Moreover, the arrangement of the cooling mechanism and the low-temperature heat sink is relatively flexible, which increases the reliability of the device. Attached Figure Description

[0056] Figure 1 This is a structural diagram of a sample preparation system according to an embodiment of this application.

[0057] Figure 2 for Figure 1 Enlarged structural diagram at point A.

[0058] Figure 3 This is a flowchart of a sample preparation method according to an embodiment of this application.

[0059] Figure 4 This is a cross-sectional view of the sample chip in the fabrication unit region according to the first embodiment of this application.

[0060] Figure 5 This is a cross-sectional view of the sample chip at the partition portion according to the first embodiment of this application.

[0061] Figure 6 This is a cross-sectional view of the sample chip in the fabrication unit region according to the second embodiment of this application.

[0062] Figure 7 This is a cross-sectional view of the sample chip at the partition portion according to the second embodiment of this application.

[0063] Figure 8 This is a cross-sectional view of the sample chip in the fabrication unit region according to the third embodiment of this application.

[0064] Figure 9 This is a cross-sectional view of the sample chip at the partition portion according to the third embodiment of this application.

[0065] Figure 10 This is a cross-sectional view of the sample chip in the fabrication unit region according to the fourth embodiment of this application.

[0066] Figure 11 This is a cross-sectional view of the sample chip at the partition portion according to the fourth embodiment of this application.

[0067] Figure 12 This is a cross-sectional view of the sample chip in the fabrication unit region according to the fifth embodiment of this application.

[0068] Figure 13 This is a cross-sectional view of the sample chip at the partition portion according to the fifth embodiment of this application.

[0069] Figure 14 for Figure 4 The diagram shows a top view of the sample chip.

[0070] Figure 15 This is a cross-sectional view of the sample chip according to the sixth embodiment of this application.

[0071] Figure 16 for Figure 15 The diagram shows a top view of the substrate in the sample chip.

[0072] Figure 17 for Figure 15 The diagram shows a top view of the first support film in the sample carrier chip.

[0073] Figure 18 for Figure 15 The diagram shows a top view of the intermediate interconnect layer in the sample chip.

[0074] Figure 19 for Figure 15 The diagram shows a top view of the second support film in the sample carrier chip.

[0075] Figure 20 for Figure 15 The diagram shows the structure of the sample chip, in which the substrate, the first support film, and the intermediate interconnect layer are stacked together.

[0076] Figure 21 for Figure 15 A bottom view of an embodiment of the substrate in the sample chip shown.

[0077] Figure 22 for Figure 15 Another embodiment of the substrate in the sample chip is shown in a bottom view structural diagram.

[0078] Figure 23 for Figure 15 A bottom view of another embodiment of the substrate in the sample chip shown.

[0079] Figure 24 for Figure 15 The cross-sectional view of the sample chip at the separator and reinforcement sections is shown.

[0080] Figure 25 This is a top view of the first support film of the sample carrier chip according to the seventh embodiment of this application.

[0081] Figure 26 This is a structural diagram of the substrate, the first support film, and the intermediate interconnect layer stacked together in the sample carrier chip of the seventh embodiment of this application.

[0082] Explanation of reference numerals in the attached figures:

[0083] 100. Sample carrier chip; 10. Substrate; 11. First cutout area; 111. Cutout area; 12. Sample inlet channel; 13. First flow channel; 14. Second flow channel; 15. Ohmic contact resistance; 16. Reinforcing part; 20. Support member; 21. Chamber; 211. Fabrication unit area; 22. Sample inlet hole; 23. First support film; 231. First observation area; 2311. First hole; 2312. First sealing film; 232. Drainage groove; 24. Second support film; 241. Second observation area; 2411. Second hole; 2412. Second sealing film; 242. 243. Second main support section; 244. Second auxiliary support section; 245. Groove; 26. Separator; 27. Support column; 28. Intermediate connecting layer; 29. ​​Window; 30. Heating electrode; 41. Support layer; 42. Second hollow section; 50. Sample inlet; 60. Heating mechanism; 70. Support platform; 71. Cooling mechanism; 72. Liquid replenishment port; 80. Low temperature heat sink; 91. Cooling braided tape; 92. Support frame; 921. Support leg; 922. Heat insulation seat; 93. Positioning frame; 94. Adapter plate; 95. Electrical connection connector plate; 96. Base. Detailed Implementation

[0084] To make the above-mentioned objectives, features, and advantages of this application more apparent and understandable, the specific embodiments of this application are described in detail below with reference to the accompanying drawings. Many specific details are set forth in the following description to provide a thorough understanding of this application. However, this application can be implemented in many other ways different from those described herein, and those skilled in the art can make similar modifications without departing from the spirit of this application. Therefore, this application is not limited to the specific embodiments disclosed below.

[0085] See Figures 1 to 4 One embodiment of this application provides a sample preparation apparatus, including: a cooling mechanism 70, a low-temperature heat sink 80, and a heat conduction mechanism. The heat conduction mechanism is connected between the cooling mechanism 70 and the low-temperature heat sink 80, and the cooling mechanism 70 cools the sample carrier unit supported on the low-temperature heat sink 80 through the heat conduction mechanism and the low-temperature heat sink 80.

[0086] The aforementioned heat conduction mechanism has a good heat conduction effect. The cooling capacity of the cooling mechanism 70 is quickly conducted to the low-temperature heat sink 80 through the heat conduction mechanism, which can accurately control the temperature of the low-temperature heat sink 80, and thus accurately control the temperature of the sample carrying unit, ensuring high sample preparation efficiency. Moreover, the arrangement of the cooling mechanism 70 and the low-temperature heat sink 80 is relatively flexible, which increases the reliability of the device.

[0087] For example, another embodiment of this application provides a sample preparation system, including the sample preparation apparatus of the above embodiment, and further including a sample carrying unit. The sample carrying unit includes a heating mechanism 50, a support stage 60, and a sample carrier chip 100. The heating mechanism 50 is used to carry the sample carrier chip 100 for heating the sample carrier chip 100. The support stage 60 is used to carry the heating mechanism 50. The sample carrier chip 100 includes a substrate 10 and a support member 20. The substrate 10 is located at the bottom of the support member 20 and is used to support the support member 20. The substrate 10 has a first cutout portion 11. The electron beam of the transmission electron microscope can pass through the first cutout portion 11 during operation. The support member 20 is connected to the substrate 10, and the support member 20 has a chamber 21 and a sample inlet 22. The inner wall of the chamber 21 is provided as a hydrophilic wall surface, and the sample inlet 22 communicates with the chamber 21.

[0088] In the above-described sample preparation system, the heating mechanism 50 is activated during use, and the sample liquid enters the chamber 21 through the inlet 22. The heating mechanism 50 prevents the sample liquid from being carried away by the cooling mechanism 70 and freezing. Furthermore, because the inner wall of the chamber 21 is hydrophilic, the sample liquid will merge at the diffusion interface under the action of the precursor film, forming liquid bridges. In other words, the sample liquid wets the entire chamber 21 under capillary action. After the sample liquid has wetted the entire chamber 21, the heating mechanism 50 stops heating, and the cooling mechanism 70 instantly removes the heat from the sample liquid, rapidly freezing the sample liquid within the chamber 21 to form an ice layer containing the sample, i.e., the sample layer. The thickness of the sample layer depends on the distance between the two opposing walls of the chamber 21 along the thickness direction of the support member 20; therefore, the thickness of the sample layer can be precisely controlled.

[0089] The low-temperature heat sink 80 is made of materials including, but not limited to, copper, copper alloys, aluminum, and their alloys. The low-temperature heat sink 80 may be a heat-conducting block, heat-conducting plate, or heat-conducting column, and possesses excellent thermal conductivity. The low-temperature heat sink 80 is connected between the cooling mechanism 70 and the support platform 60. In this way, on the one hand, it can quickly absorb the heat transferred from the support platform 60, controlling the temperature of the support platform 60 within a preset range; on the other hand, the cooling mechanism 70 conducts cold energy to the low-temperature heat sink 80, which controls the temperature within the preset range, resulting in high temperature control stability.

[0090] To improve cooling efficiency, the cooling mechanism 70 may, for example, be a liquid gas container, such as a liquid nitrogen container or a liquid ethane container. Taking a liquid nitrogen container as an example, the liquid nitrogen temperature in the container reaches -196°C, providing good cooling performance.

[0091] In one specific embodiment, the heat conduction mechanism includes a cooling braided strip 91. The cooling braided strip 91 is, for example, a soft braided strip, offering greater flexibility. To improve the cooling effect, the cooling braided strip 91 is made of at least one material, including but not limited to oxygen-free copper or graphite sheets. The graphite sheets are cooling sheets made of graphite material, possessing good thermal conductivity. The cooling braided strip 91 connects the cooling mechanism 70 to the low-temperature heat sink 80. Specifically, one end of the cooling braided strip 91 is thermally connected to the cooling mechanism 70 to absorb the cold energy from the cooling mechanism 70; the other end of the cooling braided strip 91 is thermally connected to the low-temperature heat sink 80, conducting the cold energy to the low-temperature heat sink 80. Thus, the cold energy of the liquid gas in the cooling mechanism 70 is conducted to the low-temperature heat sink 80 through the cooling braided strip 91, and then conducted to the support stage 60 and the sample chip 100 on the support stage 60, achieving rapid cooling of the sample chip 100. Thus, this application employs a solid-solid heat transfer method to conduct cooling to the sample carrier chip 100, avoiding the shear force generated on the sample after direct contact with liquids such as liquid nitrogen / liquid ethane and boiling, thus avoiding the potential damage to the sample caused by shear force. In addition, it avoids the impact of vibration generated by the boiling of liquid gas on sample loading. Furthermore, the solid-solid heat transfer method eliminates the gas film generated by liquid coolant during phase change, which is beneficial for improving the cooling rate.

[0092] Based on the aforementioned embodiments, when the cooling mechanism 70 includes a liquid gas container, the liquid gas container is provided with a liquid storage cavity. The top of the liquid storage cavity is open, allowing the liquid gas inside the liquid storage cavity to be exposed through the top of the liquid storage cavity. That is, the liquid gas container adopts an open chamber structure, which is beneficial for forming an absolutely dry nitrogen environment around the sample during sample preparation, avoiding sample contamination. Furthermore, the heat-conducting braided tape 91 can extend into the liquid storage cavity through the top of the liquid storage cavity and come into contact with the liquid gas to achieve a thermally conductive connection between the two, so that the cold energy of the liquid storage cavity can be effectively conducted to the heat-conducting braided tape 91.

[0093] For example, the other end of the cooling braided tape 91 can be fixed to the low-temperature heat sink 80, so as to be tightly combined with the low-temperature heat sink 80 to realize the transfer of cold energy; of course, the other end of the cooling braided tape 91 can also be attached to the low-temperature heat sink 80 to realize the transfer of cold energy to the low-temperature heat sink 80.

[0094] For example, the liquid gas container includes an inner layer, a middle layer, and an outer layer arranged sequentially from the inside out. The inner layer is made of PCTFE material that can withstand low temperatures. The middle layer is made of EPS foam material, which has vibration damping and heat insulation effects. The outer layer is made of plastic and serves a protective function.

[0095] For example, the liquid gas container is provided with a replenishment port 71, through which liquid gas can be added to the storage cavity of the liquid gas container in a timely manner. The liquid gas container is also provided with a liquid level sensor 72. The liquid level sensor 72 is used to detect the liquid level height of the liquid gas in the storage cavity. A control valve is provided at the replenishment port 71. When the liquid level sensor 72 detects that the liquid level is lower than a first preset liquid level height, the control valve opens the replenishment port 71, and liquid gas is automatically replenished through the replenishment port 71; when the liquid level sensor 72 detects that the liquid level is higher than a second preset liquid level height, for example, when it detects that the liquid gas has filled the storage cavity, the control valve closes the replenishment port 71, and stops the replenishment of liquid gas.

[0096] Based on the aforementioned embodiments, the sample preparation apparatus further includes a support frame 92. The support frame 92 is arranged adjacent to the liquid gas container. A cryogenic heat sink 80 is installed on top of the support frame 92. Specifically, both the support frame 92 and the liquid gas container are fixedly mounted on the support platform. The height of the cryogenic heat sink 80 is higher than that of the liquid gas container, so that the sample carrier chip 100 is located in the area above the top of the liquid gas container. One end of the cooling braided tape 91 extends downward from the top of the liquid storage cavity of the liquid gas container into the liquid storage cavity, and the other end of the cooling braided tape 91 is located outside the liquid gas container and is thermally connected to the cryogenic heat sink 80. The top opening of the liquid storage cavity is open, which allows the air in the area above the top of the liquid gas container to be dry, providing a dry environment for the sample carrier chip 100 and avoiding sample contamination.

[0097] Of course, as an alternative, the height of the cryogenic heat sink 80 can also be lower than that of the liquid gas container, and no specific restrictions are imposed here.

[0098] Please see Figure 1 and Figure 2Based on the aforementioned embodiments, the support frame 92 includes legs 921 and a heat insulation base 922. The legs 921 are connected to the heat insulation base 922. The legs 921 support the heat insulation base 922, ensuring its stability. The legs 921 may include, but are not limited to, support columns or support plates. Optionally, there are two legs 921 arranged side-by-side with a gap between them, and the heat insulation base 922 is connected between the two legs 921. This provides stable support for the heat insulation base 922 and allows its bottom to be suspended, reducing contact with other components and providing better heat insulation. The legs 921 and the heat insulation base 922 are made of different materials. The legs 921 and the heat insulation base 922 are connected and fixed by fasteners such as pins, rivets, and screws. A low-temperature heat sink 80 is installed on the heat insulation base 922. The heat insulation base 922 provides heat insulation, reducing energy loss of the low-temperature heat sink 80. The heat insulation seat 922 can stably support the low-temperature heat sink 80. Part of the low-temperature heat sink 80 is made of the heat insulation seat 922, and the other part is suspended.

[0099] To facilitate various operations such as sample loading and observation of the sample carrier chip 100, based on the aforementioned embodiment, at least a portion of the support stage 60 is disposed on the low-temperature heat sink 80, thereby contacting the low-temperature heat sink 80 and allowing the low-temperature heat sink 80 to conduct heat to the support stage 60, thus achieving a cooling effect. Another portion of the support stage 60 extends beyond the low-temperature heat sink 80 and is suspended, so that the projection of the sample carrier chip 100 along the thickness direction is offset from the low-temperature heat sink 80. That is, the suspended portion of the support stage 60 is used to support and carry the sample carrier chip 100, providing ample space above and below the sample carrier chip 100 to integrate sample loading devices, observation devices, positioning devices, or thickness measuring devices. These devices can then be arranged above and below the suspended portion of the support stage 60, enabling various operations on the sample carrier chip 100.

[0100] Please see Figure 2 Based on the aforementioned embodiments, the sample preparation device further includes a positioning frame 93. A portion of the positioning frame 93 is connected to the low-temperature heat sink 80, and the other portion extends beyond the low-temperature heat sink 80 and is suspended in mid-air. The positioning frame 93 is used to support and position the support platform 60. Thus, the positioning frame 93 serves to support and position the support platform 60, improving the accuracy of the loading position of the support platform 60 and enhancing the stability of the support platform 60 and the sample-carrying chip 100. Optionally, the positioning frame 93 is adapted to the shape of the support platform 60 to stably support it.

[0101] For example, the sample carrying unit also includes a clamping component. The clamping component includes, but is not limited to, clamping screws, clamping bolts, clamping plates, etc., and is connected to the support stage 60. The clamping component is used to press and fix the sample chip 100 and the heating mechanism 50 firmly onto the support stage 60. Thus, under the clamping force of the clamping component, good thermal contact is formed between the sample chip 100, the heating mechanism 50, and the support stage 60, and the heat from the heating mechanism 50 is effectively conducted to the sample chip 100. When the heating mechanism 50 stops working, the support stage 60 can conduct the cooling energy from the cooling mechanism 70 to the sample chip 100, causing the sample inside the sample chip 100 to freeze rapidly.

[0102] Please see Figure 1 and Figure 2 Based on the aforementioned embodiments, the sample preparation device further includes a base 96. A cooling mechanism 70 is mounted on the base 96. Furthermore, a support frame 92 is mounted on the base 96.

[0103] For example, the heating mechanism 50 includes a heating chip. The heating chip is stacked and cooperates with the sample carrier chip 100 to conduct heat to the sample carrier chip 100. The sample carrier unit also includes an adapter plate 94. The heating chip is electrically connected to the adapter plate 94, and the heating chip is stacked on the adapter plate 94. The heating chip is fixed to the adapter plate 94. For example, the heating chip forms an electrical connection and thermal contact with the metal leads of the adapter plate 94 by bonding. Of course, the heating chip can also be connected to the adapter plate 94 in other ways, without particular limitation. The adapter plate 94 is disposed on the carrier stage 60. Specifically, the adapter plate 94 is packaged and fixed to the carrier stage 60, so that the adapter plate 94, the carrier stage 60, the heating mechanism 50 and the sample carrier chip 100 are combined into a whole, which can be easily grasped by the robot and placed on the low-temperature heat sink 80.

[0104] In some specific embodiments, the heating chip and the adapter plate 94 are connected by adhesive bonding. During actual assembly, the adapter plate 94 is first placed on a fixture, which is vacuumed to hold the adapter plate 94 in place to prevent movement; then the positioning component is placed on the fixture, and the heating chip is positioned at the designated location on the adapter plate 94 via the positioning component; next, the pressure component presses the heating chip together, and the heating chip is bonded to the adapter plate 94 by heating and pressurizing, so that the heating chip and the adapter plate 94 are fixedly connected together.

[0105] Furthermore, after the adapter plate 94 is fixedly connected to the heating chip, the entire assembly formed by the adapter plate 94 and the heating chip is placed in the preset position of the carrier platform 60; so that the adapter plate 94, the heating chip and the carrier platform 60 are placed together in the fixing component; the external hot press applies pressure to the adapter plate 94 through the pressure component, so that the adapter plate 94 and the carrier platform 60 are bonded and fixed to each other, thereby realizing the encapsulation and fixation of the adapter plate 94 on the carrier platform 60.

[0106] In some embodiments, the heating mechanism 50 is provided with a positioning groove for positioning the sample chip 100, and the sample chip 100 is disposed in the positioning groove and bonded and fixed to the heating mechanism 50. In this way, the sample chip 100 is stably disposed on the heating mechanism 50, thereby ensuring the heat conduction effect.

[0107] Based on the aforementioned embodiment, the heating mechanism 50 has a first cutout, and the support stage 60 has a second cutout. Both the first and second cutouts are opposite to the sample carrier chip 100 along its thickness direction. Thus, when a transmission electron microscope observes a frozen sample on the sample carrier chip 100, electrons can pass through the second and first cutouts. This avoids obstruction and interference of electrons by the heating mechanism 50 and the support stage 60.

[0108] For example, the sample carrier unit also includes an electrical connection connector plate 95. The adapter plate 94 is electrically connected to the electrical connection connector plate 95, which is used for electrical connection to the controller. Under the control of the controller, the heating chip can be controlled to operate or stop operating, thereby controlling the temperature of the heating chip and, consequently, accurately controlling the temperature of the sample carrier chip 100. During sample loading, to prevent the sample liquid from freezing and becoming unloadable, the temperature of the sample carrier chip 100 is controlled at, for example, above 4°C by the heating chip. After the sample carrier chip 100 has finished loading, the heating chip is de-energized, and the sample inside the sample carrier chip 100 will rapidly freeze.

[0109] Specifically, the electrical connection connector plate 95 is mounted on the fixed frame. The electrical connection connector plate 95 is equivalent to a connector for electrical connection with the controller.

[0110] Please see Figure 3 , Figure 3 A flowchart of a sample preparation method according to an embodiment of this application is shown. An embodiment of this application provides a sample preparation method using a sample preparation system, including:

[0111] Step S100: Turn on the heating mechanism 50 to heat the sample carrier chip 100 and control the temperature of the sample carrier chip 100 to be higher than the first preset temperature.

[0112] Optionally, the first preset temperature is above 4°C, specifically such as 4°C, 6°C, 10°C or 20°C, etc. There is no restriction here, as long as it can prevent the sample liquid temperature from being too low and causing freezing.

[0113] Step S200: The sample liquid is added into the chamber 21 through the inlet port 22. The sample liquid spontaneously wets the chamber 21 through capillary action.

[0114] Step S300: After the sample liquid wets the entire chamber 21, the heating mechanism 50 stops working, and the cooling mechanism 70 lowers the temperature of the sample carrier chip 100 to below the second preset temperature, so that the sample liquid in the sample carrier chip 100 freezes rapidly.

[0115] Optionally, the second preset temperature may include, but is not limited to, less than 0°C, such as -5°C, -10°C, -20°C, -50°C, etc. The specific temperature can be flexibly adjusted and set according to actual needs, as long as the sample liquid inside the sample carrier chip 100 can be quickly frozen.

[0116] The above-described sample preparation method corresponds to the sample preparation system, and therefore the beneficial effects are the same as those of the sample preparation system, so they will not be repeated here.

[0117] Please refer to the following: Figure 4 Optionally, the injection port can be located on any side of the chamber along the thickness direction of the support, that is, it can be located on the side of the chamber away from the substrate or on the side of the chamber facing the substrate, without limitation. The number of injection ports 22 can be, for example, one, two, three, four or other numbers, without limitation.

[0118] In one embodiment, the sample inlet 22 is located on a portion of the support 20 exposed to the external environment. Specifically, the sample inlet 22 is, for example, located on the side of the support 20 facing away from the substrate 10, such as... Figure 4 As shown; of course, the sample inlet 22 can also be, for example, located on the side of the substrate 10 facing the substrate 10 and corresponding to the area of ​​the first cutout 11. When adding sample liquid, the sample liquid can be directly added to the sample inlet 22 and enter the chamber 21 through the sample inlet 22. Since the inner wall of the chamber 21 is designed as a hydrophilic wall, the sample liquid will merge at the diffusion interface under the action of the precursor film and form a liquid bridge, that is, the sample liquid wets the entire chamber 21 under the action of capillary force. After the sample liquid wets the entire chamber 21, the sample liquid in the chamber 21 is rapidly frozen to form an ice layer containing the sample, that is, the sample layer.

[0119] Of course, in other embodiments, the injection port 22 may also be located on a part of the support 20 that is not exposed to the external environment, such as... Figures 15 to 20 As shown. Specifically, the sample inlet 22 is, for example, located on the side of the support 20 facing the substrate 10 and covered by the substrate 10 along the thickness direction of the support 20. The sample liquid is not directly added into the sample inlet 22. How it is added into the sample inlet 22 will be explained in detail in the embodiments below, and will not be repeated here.

[0120] Please refer to the following: Figure 4Optionally, the support member 20 includes a first support film 23 and a second support film 24 disposed at a distance from each other. The first support film 23 is connected to the substrate 10, and the second support film 24 is located on the side of the first support film 23 opposite to the substrate 10 and cooperates with the first support film 23 to form a chamber 21. The first support film 23 is provided with a first observation portion 231. The second support film 24 is provided with a second observation portion 241, and the second observation portion 241 is arranged opposite to the first observation portion 231.

[0121] Based on the aforementioned embodiments, the projections of the first observation section 231 and the second observation section 241 along the thickness direction of the support member 20 are both located within the contour range of the first hollow section 11. When the electron beam passes through the observation section and through the support member 20, the electron beam can acquire sample information within the chamber 21 of the support member 20. After acquiring the sample information, the electron beam continues to pass through the sample carrier chip 100 along the first hollow section 11 and propagates to the transmission electron microscope's electron beam receiving device to acquire sample information, ultimately achieving imaging.

[0122] In the aforementioned sample carrier chip 100, the thickness of the sample layer depends on the distance between the two walls of the chamber 21 that are disposed opposite to each other along the thickness direction of the support member 20. Therefore, the thickness of the sample layer can be precisely controlled.

[0123] In addition, the first observation section 231, the second observation section 241 and the first hollow section 11 facilitate the passage of the electron beam of the transmission electron microscope, which can meet the data collection needs and match the pipeline-style cryo-electron microscope data collection, thereby improving the collection efficiency of cryo-electron microscope image data.

[0124] Furthermore, the substrate 10 is used for mounting onto the sample transfer device of a transmission electron microscope or onto a grid. In other words, the sample carrier chip 100 can be used alone or in combination with a grid. The sample transfer device refers to the device used to place the sample into the electron microscope, such as a sample holder or a grid transfer device. The sample carrier chip 100 can directly use sample transfer devices in related technologies, which will not be described in detail here.

[0125] The substrate 10 ensures the flatness of the support 20, allowing the cryo-electron microscope to meet observation requirements with a single focusing during data collection, saving time spent on the focusing process and improving efficiency. Furthermore, the sample carrier chip 100 facilitates mass production, eliminating the need for manual application of the support film to the carrier mesh, thus improving processing efficiency.

[0126] In this application, the substrate 10 includes, but is not limited to, a silicon wafer. The silicon wafer includes, but is not limited to, monocrystalline silicon, polycrystalline silicon, or silicides, etc., and is not limited thereto. Silicides include, for example, silicon oxide or silicon nitride. The conductivity of the substrate 10 is typically weaker than that of the support 20. If the accumulated charge first moves from the support 20 to the substrate 10 and then is discharged outwards from the substrate 10, it will undoubtedly reduce the charge discharge rate of the sample carrier chip 100. Therefore, in this embodiment, the support 20, for example, directly contacts an external conductive element, thereby increasing the charge discharge rate and further reducing the risk of sample quality degradation or sample damage due to charge accumulation.

[0127] In some embodiments, please refer to Figure 4 The sample carrier chip 100 also includes an ohmic contact resistor 15. The ohmic contact resistor 15 is, but is not limited to, made of metal materials such as aluminum or titanium. The ohmic contact resistor 15 is electrically connected to at least one of the substrate 10 and the support member 20. Specifically, two ohmic contact resistors 15 are provided, arranged at intervals, and electrically connected to the positive and negative terminals of external conductive members, respectively, thereby enabling charge discharge from the sample carrier chip 100.

[0128] Specifically, the observation areas of the first observation section 231 and the second observation section 241 are sufficient to meet the data collection requirements of the transmission electron microscope, and are matched with automated cryogenic data collection to improve the efficiency of electron microscope image data collection.

[0129] Specifically, in conventional transmission electron microscopy (TEM), approximately one-third of the machine time is spent on focusing during data collection. The flatness of the support 20 in this application is, for example, better than 1 μm, which allows the TEM to focus only once during data collection, saving the time spent on focusing.

[0130] Please see Figure 4 Based on the aforementioned embodiments, there are, for example, two injection ports 22. To facilitate sample addition, the two injection ports 22 are arranged, for example, on the same side of the chamber 21. More specifically, to improve sample addition efficiency and the uniformity of wetting of the chamber 21, the two injection ports 22 are arranged, for example, at two diagonal locations on the same side of the chamber 21.

[0131] In this embodiment, the sample carrier chip 100 is not limited to being fabricated using MEMS technology. MEMS technology can achieve mass production of sample carrier chips 100, with reliable technology and high yield.

[0132] For example, the contact angle of the inner wall of chamber 21 may include, but is not limited to, 85°, 80°, 75°, 70°, 65°, 60°, 55°, 50°, 45°, 40°, 35°, 30°, 25°, 20°, 15°, 10°, 9°, 8°, 7°, 6°, 5°, 4° or 3°, etc. The specific angle can be flexibly adjusted and set according to actual needs, and is not limited here.

[0133] Specifically, the better the hydrophilicity of the inner wall of chamber 21, the more conducive it is to wetting the entire surface of the inner wall of chamber 21 and the shorter the wetting time. Therefore, in this embodiment, the contact angle of the inner wall of chamber 21 is preferably less than 30°, more preferably less than 10°, further less than 5°, or even less than 3°.

[0134] For example, in this embodiment, the inner wall of the chamber 21 is a superhydrophilic wall, that is, the contact angle of the inner wall of the chamber 21 is less than 5°.

[0135] In order to make the inner wall of the chamber 21 hydrophilic, for example, the inner wall of the chamber 21 may be subjected to treatments including but not limited to plasma hydration or ALD titanium dioxide modification. Of course, other treatments may be performed on the inner wall of the chamber 21 to make the inner wall of the chamber 21 hydrophilic, which is not limited here.

[0136] For example, at least one side of the inner wall of chamber 21 is entirely hydrophilic, while the degree of hydrophilicity of the other side of the inner wall of chamber 21 is not limited. Specifically, all parts of the inner walls of opposite sides of chamber 21 along the thickness direction of support 20 are hydrophilic. In this embodiment, all parts of the opposing walls of the first support membrane 23 and the second support membrane 24 are hydrophilic. Thus, the sample liquid can wet the entire inner wall of chamber 21.

[0137] For example, the outer wall of the support 20 is designed to be hydrophobic, meaning the contact angle is greater than 90°. The outer wall of the support 20 refers to the wall facing the external environment, or the wall away from its geometric center. Specifically, in this embodiment, the outer wall of the support 20 refers to the side of the support 20 away from the substrate 10. More specifically, the outer wall of the support 20 is designed to be superhydrophobic, meaning the contact angle is greater than 150°. This effectively prevents the sample liquid inside the chamber 21 from overflowing from the inlet 22 or observation section onto the outer wall of the support 20. This also facilitates the complete entry of the sample liquid into the chamber 21 during sample addition, filling the entire chamber 21 under capillary action, thereby enabling precise control of the sample layer thickness. Furthermore, it results in good sample uniformity and a high sample yield.

[0138] For example, the contact angle of the outer wall of the support member 20 may include, but is not limited to, 92°, 95°, 98°, 100°, 105°, 110°, 115°, 120°, 125°, 130°, 135°, 140°, 145°, 150°, 155°, 160°, 165°, 170° or 175°, etc. The specific angle can be flexibly adjusted and set according to actual needs, and is not limited here.

[0139] Please see Figure 4 For example, the distance between the inner walls of opposite sides of the chamber 21 along the thickness direction of the support 20 is S, where 20nm ≤ S ≤ 150nm. That is, the distance between the first support film 23 and the second support film 24 is S. The thickness of the sample layer prepared by the sample carrier chip 100 is equal to the distance S, corresponding to a thickness between 20nm and 300nm, thus meeting the requirements. Specifically, S includes, but is not limited to, 20nm, 30nm, 40nm, 50nm, 60nm, 70nm, 80nm, 90nm, 100nm, 110nm, 120nm, 130nm, 140nm, 150nm, 200nm, 250nm, 280nm, or 300nm, etc., which can be flexibly adjusted and set according to actual needs, and are not limited here.

[0140] The thickness of the sample layer can affect the test results. Therefore, in order to enrich the test data, it is necessary to prepare sample layers of various thicknesses and test the sample layers of different thicknesses.

[0141] In this embodiment, to improve the preparation and detection efficiency of sample layers of different thicknesses, multiple sample layers of different thicknesses are prepared using the same sample carrier chip 100. The number of sample layers of different thicknesses that can be prepared by the same sample carrier chip 100 is, for example, 5, 10, 20, or 100, etc., and is not limited here. Compared to being able to prepare only one type of sample layer, both preparation and detection efficiency are improved.

[0142] Please see Figure 4 and Figure 14Specifically, the chamber 21 is provided with multiple preparation unit areas 211. Each preparation unit area 211 is used to prepare a sample layer, and each preparation unit area 211 corresponds to preparing a sample layer of a certain thickness. The sample layer thicknesses prepared by each preparation unit area 211 may be exactly the same, not exactly the same, or completely different; this is not limited and can be set according to actual needs. In one embodiment, the sample layer thicknesses prepared by each preparation unit area 211 are completely different, allowing the sample carrier chip 100 to prepare a variety of sample layers of different thicknesses to meet detection requirements. That is, the spacing between the opposite inner walls of each preparation unit area 211 along the thickness direction of the support member 20 is different.

[0143] For example, for the preparation unit region 211, the inner walls of the opposite sides of the preparation unit region 211 along the thickness direction of the support member 20 are arranged parallel to each other, so that the sample layer prepared by the preparation unit region 211 has high uniformity of thickness, controllable sample layer thickness, and high sample yield.

[0144] The arrangement of multiple preparation unit regions 211 within the chamber 21 can take many forms, such as being arranged sequentially along a straight line, sequentially along a curve, arranged in a matrix, or arranged in other regular or irregular ways. No particular limit is imposed here, and the arrangement can be flexibly selected according to actual needs.

[0145] In this embodiment, we will take the arrangement of multiple fabrication unit regions 211 in a straight line as an example.

[0146] For example, multiple fabrication unit regions 211 are arranged sequentially in a straight line, and the arrangement direction is perpendicular to the thickness direction of the support 20. Specifically, the multiple fabrication unit regions 211 are, for example, along the length direction of the support 20 (e.g., Figure 14 As shown by arrow Y in the diagram, they can be arranged sequentially along the width direction of the support member 20 (e.g., ...). Figure 14 Arranged sequentially on the arrow X shown in the image.

[0147] Based on the aforementioned embodiments, along the arrangement direction of the preparation unit regions 211, the spacing between the inner walls of the opposite sides of the preparation unit regions 211 along the thickness direction of the support member 20 tends to increase or decrease. Thus, the thickness of the prepared sample layer tends to increase or decrease along its arrangement direction, which can help improve the accuracy of the detection results of each sample layer. Specifically, for any two adjacent preparation unit regions 211 along the arrangement direction, the difference in the spacing between the inner walls of the opposite sides of the preparation unit regions 211 along the thickness direction of the support member 20 is, but not limited to, 5 nm to 20 nm. Therefore, the difference in the thickness of the sample layer prepared by two adjacent preparation unit regions 211 is, but not limited to, 5 nm to 20 nm, specifically, for example, 5 nm, 8 nm, 10 nm, 12 nm, 15 nm, 18 nm, or 20 nm.

[0148] Of course, as some alternatives, the spacing between the inner walls of the two opposite sides of the preparation unit region 211 along the thickness direction of the support member 20 can also be distributed in an irregular manner along the arrangement direction of the preparation unit region 211, and is not limited here.

[0149] Please see Figure 4 , Figure 5 and Figure 14 For example, the support member 20 includes a plurality of partitions 25 disposed within a chamber 21. The inner walls of opposite sides of the chamber 21 along the thickness direction of the support member 20 are connected to the partitions 25. Each partition 25 is correspondingly disposed between two adjacent preparation unit regions 211 along the arrangement direction. Thus, on the one hand, the partitions 25 separate two adjacent preparation unit regions 211, reducing mutual interference between them and improving the detection accuracy of each sample layer. On the other hand, the partitions 25 increase the structural strength of the support member 20. Especially for a support member 20 comprising two opposing amorphous films, the partitions 25 provide support for the amorphous films, preventing them from adhering to each other, which helps ensure the thickness uniformity of the sample layers and improves the sample preparation success rate.

[0150] For example, there are multiple first observation sections 231 and multiple second observation sections 241. The number of each first observation section 231 and the number of each second observation section 241 are the same as the number of preparation unit regions 211. Each first observation section 231 is provided corresponding to each preparation unit region 211, and each second observation section 241 is provided corresponding to each preparation unit region 211.

[0151] For example, the first observation section 231 and the second observation section 241 have the same shape and size, and are aligned with each other along the thickness direction of the support member 20. Since the shape and size of the second observation section 241 are the same as those of the first observation section 231, the following description will focus on the first observation section 231. The first observation section 231 is elongated, for example, linearly extended, specifically as a slit. That is, the width of the first observation section 231 is small and the length is large. The sample chip 100 only needs to move in the length direction to ensure comprehensive observation, which is beneficial to the motion control of the sample chip 100 and can also avoid interference between the sample chip 100 and the pole piece of the transmission electron microscope.

[0152] Specifically, the transmission electron microscope (TEM) is equipped with pole pieces for emitting and receiving electron beams. The electron beam emitted from the pole pieces can pass through the first observation section 231 and the second observation section 241 and be focused on the preparation unit region 211, acquiring information about the sample within the preparation unit region 211. This information is then transmitted to the pole pieces, thus enabling sample observation. In actual implementation, as the sample carrier chip 100 moves along the length of the first observation section 231, the electron beam continuously acquires information about the sample, ensuring comprehensive observation.

[0153] Therefore, the sample carrier chip 100 can perform linear, pipelined cryogenic data collection, improving the efficiency of electron microscope image data collection.

[0154] Please see Figure 4 and Figure 14 In some examples, the width of the first observation section 231 is W, which includes, but is not limited to, 0.6μm-2.5μm, and can be flexibly adjusted and set according to actual needs. Specifically, the width W of the first observation section 231 can be set to 0.6μm, 0.8μm, 0.9μm, 1μm, 1.2μm, 1.5μm, 1.6μm, 1.7μm, 2μm, or 2.5μm, etc. It can be seen that the area of ​​the first observation section 231 is approximately 10 times the area required for data collection, which facilitates matching with pipelined cryogenic data collection while meeting data collection requirements.

[0155] Please see Figure 4 For example, the first observation section 231 includes a first hole 2311 penetrating the first support membrane 23, and the second observation section 241 includes a second hole 2411 penetrating the second support membrane 24. Thus, when the transmission electron microscope is in operation, the electron beam emitted from the pole piece can pass through the first observation section 231 and the second observation section 241 and be focused on the preparation unit region 211, acquiring information about the sample within the preparation unit region 211, and then transmitting the sample information to the pole piece, thereby achieving sample observation.

[0156] Both the first support film 23 and the second support film 24 are fabricated using MEMS technology, resulting in a high yield. For sample introduction, the injection port 22 can be located on either the first support film 23 or the second support film 24; this is not a limitation. However, to facilitate sample addition, in this embodiment, the injection port 22 is specifically located on the second support film 24.

[0157] For example, the first support film 23 includes a first main support portion and a first auxiliary support portion. The first auxiliary support portion is connected to the first main support portion, and a first hole 2311 is disposed in the first auxiliary support portion. Specifically, the number of first auxiliary support portions is the same as the number of fabrication unit regions 211, and each first auxiliary support portion is correspondingly disposed to each fabrication unit region 211. The area size of the first auxiliary support portion is designed according to the area size of the fabrication unit region 211. The thickness of the first auxiliary support portion is less than the thickness of the first main support portion, and the first auxiliary support portion and the first main support portion cooperate to form a stepped shape. The first main support portion serves as the main structure of the first support film 23, and its area size and thickness are both greater than those of the first auxiliary support portion. In this way, the excessive thickness of the first auxiliary support portion can be avoided, which would cause the gas-liquid interface charge to have an adverse effect on imaging, thus improving the imaging quality; in addition, the larger thickness of the first main support portion can ensure the structural strength of the first support film 23.

[0158] Please see Figure 4 For example, the second support film 24 includes a second main support portion 242 and a second auxiliary support portion 243. The second auxiliary support portion 243 is connected to the second main support portion 242, and a second hole 2411 is disposed in the second auxiliary support portion 243. Specifically, the number of second auxiliary support portions 243 is the same as the number of preparation unit regions 211, and each second auxiliary support portion 243 is correspondingly disposed to each preparation unit region 211. The area size of the second auxiliary support portion 243 is designed according to the area size of the preparation unit region 211. Each second auxiliary support portion 243 corresponds to each first auxiliary support portion and is disposed at a relative interval. The area where the corresponding first auxiliary support portions and second auxiliary support portions 243 cooperate with each other is the preparation unit region 211.

[0159] The thickness of the second auxiliary support 243 is less than the thickness of the second main support 242. The second auxiliary support 243 and the second main support 242 cooperate to form a stepped shape, specifically, for example, a groove 244. Figure 4 or Figure 19As shown, the second main support portion 242 serves as the main structure of the second support film 24. The area and thickness of the second main support portion 242 are both larger than those of the second auxiliary support portion 243. This avoids the adverse effects of gas-liquid interface charge on imaging caused by excessive thickness of the second auxiliary support portion, thus improving imaging quality. Furthermore, the greater thickness of the second main support portion 242 ensures the structural strength of the second support film 24.

[0160] Of course, in some embodiments, the thickness of the first auxiliary support portion can also be consistent with the thickness of the first main support portion, such as... Figure 4 As shown; similarly, the thickness of the second auxiliary support 243 can also be consistent with the thickness of the second main support 242.

[0161] In some embodiments, the thickness of the first support film 23 includes, but is not limited to, 40nm to 1000nm, specifically, 40nm, 60nm, 80nm, 100nm, 200nm, 300nm, 400nm, 500nm, 600nm, 700nm, 800nm, 900nm or 1000nm, etc. The specific thickness can be flexibly selected and set according to actual needs, and is not limited here.

[0162] In some embodiments, the thickness of the second support film 24 includes, but is not limited to, 40nm to 2000nm, specifically, 40nm, 60nm, 80nm, 100nm, 200nm, 300nm, 400nm, 500nm, 600nm, 700nm, 800nm, 900nm, 1000nm, 1300nm, 1600nm, or 2000nm, etc. The specific thickness can be flexibly selected and set according to actual needs, and is not limited here.

[0163] Based on the aforementioned embodiments, the first main support portion is arranged circumferentially around the first auxiliary support portion. The outer periphery of the first auxiliary support portion is supported by the first main support portion, resulting in high stability. Furthermore, the first auxiliary support portion and the first main support portion are an integrated structure.

[0164] Based on the aforementioned embodiment, the second main support portion 242 is arranged circumferentially around the second auxiliary support portion 243. The outer periphery of the second auxiliary support portion 243 is supported by the second main support portion 242, resulting in high stability. Furthermore, the second auxiliary support portion 243 and the second main support portion 242 are an integrated structure.

[0165] Please compare Figure 4 and Figure 6 In contrast Figure 4The difference in the structure shown is that, in some embodiments, the first observation section 231 further includes a first sealing membrane 2312 connected to the first support membrane 23. The first sealing membrane 2312 is used to seal the first hole 2311. Optionally, the first sealing membrane 2312 may be located on the inner or outer side of the first support membrane 23 or arranged within the first hole 2311. The specific arrangement of the first sealing membrane 2312 is not limited and can be flexibly adjusted and set according to actual needs, as long as it can seal the first hole 2311 and prevent the sample liquid in the chamber 21 from overflowing from the first hole 2311, thus eliminating the influence of gas-liquid interface charge on imaging. Furthermore, the first sealing membrane 2312 may include, but is not limited to, graphene or other single-atom or multi-atom materials. In this way, the first sealing membrane 2312 is transparent under electron beam irradiation, and the electron beam emitted during transmission electron microscopy can pass through the first sealing membrane 2312 normally to obtain sample information without affecting the observation of the sample.

[0166] Please see Figure 10 or Figure 12 Similarly, the second observation section 241 also includes a second sealing membrane 2412 connected to the second support membrane 24. The second sealing membrane 2412 is used to seal the second hole 2411. Optionally, the second sealing membrane 2412 may be located on the inner or outer side of the second support membrane 24 or arranged inside the second hole 2411. The specific arrangement of the second sealing membrane 2412 is not limited and can be flexibly adjusted and set according to actual needs, as long as it can seal the second hole 2411 and prevent the sample liquid in the chamber 21 from overflowing from the second hole 2411, thus eliminating the adverse effects of gas-liquid interface charge on imaging. Furthermore, the second sealing membrane 2412 may include, but is not limited to, graphene or other single-atom or multi-atom materials. In this way, the second sealing membrane 2412 is transparent under electron beam irradiation, and the electron beam emitted during transmission electron microscopy can pass through the second sealing membrane 2412 normally to obtain sample information without affecting the observation of the sample.

[0167] Please continue reading. Figure 10 or Figure 12 Since neither the first observation section 231 nor the second observation section 241 has gas-liquid interface charge, based on the aforementioned embodiment, the thickness of each part of the first support film 23 is consistent, and the thickness of each part of the second support film 24 is consistent. Thus, compared to setting both the first support film 23 and the second support film 24 to have inconsistent thicknesses, it is easier to obtain a first support film 23 and a second support film 24 with consistent thicknesses during the deposition process, thereby improving both production efficiency and quality.

[0168] Of course, it should be noted that in order to simplify the manufacturing process, reduce production difficulty, and improve the production efficiency of the sample chip 100, please refer to [link / reference needed]. Figure 4 or Figure 8 The first observation section 231 may consist only of the first hole 2311, without the need for the first sealing membrane 2312 to block the first hole 2311. Similarly, the second observation section 241 may consist only of the second hole 2411, without the need for the second sealing membrane 2412 to block the second hole 2411. Furthermore, to reduce the influence of gas-liquid interface charge on imaging, the thickness of the first support membrane 23 and the second support membrane 24 can be reduced as much as possible. Alternatively, the first support membrane 23 can be divided into a first main support section and a first auxiliary support section, reducing the thickness of the first auxiliary support section, and the second support membrane 24 can be divided into a second main support section 242 and a second auxiliary support section 243, reducing the thickness of the second auxiliary support section.

[0169] Because the sample layer is relatively thin, chamber 21 is prone to adhesion due to capillary forces. Therefore, please refer to [the relevant documentation / reference]. Figure 4 , Figure 6 , Figure 8 , Figure 10 or Figure 12 To prevent the chamber 21 from sticking together due to capillary force during sample injection, the support member 20 further includes a support column 26, building upon the aforementioned embodiment. The support column 26 connects the first support membrane 23 and the second support membrane 24. Thus, the support column 26 provides support, effectively preventing the first support membrane 23 and the second support membrane 24 from sticking together, thereby ensuring that the sample liquid wets the entire chamber 21 under capillary force.

[0170] In one specific embodiment, multiple fabrication unit regions 211 are along the length direction of the support 20 (e.g., Figure 14 The components are arranged sequentially along the path indicated by arrow Y in the diagram. Furthermore, the preparation unit area 211 is located in the middle of the support 20 along its width. Additionally, there are two injection ports 22, each located at a opposite corner of the second support membrane 24. When the sample liquid is simultaneously injected through the two injection ports 22, the sample liquid, under the action of capillary force, first wets the opposite sides of the chamber 21 along the width of the support 20, and then wets each preparation unit area, forming liquid bridges.

[0171] In addition, multiple support pillars 26 are provided on both opposite sides of the preparation unit area 211 along the width direction of the support member 20. Optionally, the multiple support pillars 26 on the same side of the preparation unit area 211 form one or more rows, each row including at least two support pillars 26, and arranged sequentially at intervals along the length direction of the support member 20, for example. Specifically, each row of support pillars 26 is arranged sequentially at equal intervals along the length direction of the support member 20. In this way, the support pillars 26 provide good support stability for the second support film 24, effectively preventing adhesion defects between the second support film 24 and the first support film 23, and ensuring the uniformity of the thickness of the sample layer prepared in each preparation unit area 211, thereby improving the sample preparation yield.

[0172] Specifically, each support column 26 is connected between the first main support section and the second main support section 242. In this way, the support columns 26 avoid the fabrication unit area 211, thus preventing any adverse effects on imaging observation.

[0173] Please see Figure 5 , Figure 7 , Figure 9 , Figure 11 or Figure 13 In some embodiments, the support pillar 26 and the partition 25 are integrally formed with, for example, the first support film 23 and / or the second support film 24. During the fabrication process, the chamber 21, the first hole 2311, the second hole 2411, and the sample inlet 22 are formed by photolithography, etching, and other processes on the support member 20, while the unetched parts of the support member 20 are correspondingly formed with the first support film 23, the second support film 24, the support pillar 26, and the partition 25.

[0174] Of course, in some alternative solutions, the first support film 23, the support pillar 26, the separator 25 and the second support film 24 can also be processed on the substrate 10 respectively. The specific implementation is not limited here and can be set and adjusted according to actual needs.

[0175] The materials of the first support film 23, the second support film 24, the support pillar 26, and the partition 25 can all be the same, for example, all of which can be made of amorphous film materials, such as silicon nitride or silicon carbide. Of course, the materials of the first support film 23, the second support film 24, the support pillar 26, and the partition 25 can also be different, and the specific materials can be set according to actual needs, which is not limited here.

[0176] Please see Figure 4 and Figure 6For example, the sample carrier chip 100 also includes a heating electrode 30. The heating electrode 30 is connected to the side of the support 20 facing away from the substrate 10. Thus, before and during sample liquid injection, the heating electrode 30 is energized to heat the support 20, ensuring that the temperature inside the chamber 21 is, for example, greater than 0°C. This prevents the sample liquid from freezing inside the chamber 21 due to insufficient temperature, thus ensuring that the sample liquid entering the chamber 21 remains liquid and wets the entire chamber 21. Once the sample liquid has wetted the entire chamber 21, the heating electrode 30 is de-energized. The sample carrier chip 100, for example, comes into contact with a liquid nitrogen system, causing its temperature to drop rapidly, allowing the sample liquid inside the chamber 21 to quickly freeze and form a sample layer. This allows for precise control of the sample layer thickness, thereby improving imaging performance.

[0177] For example, the heating electrodes 30 are arranged on opposite sides of multiple preparation unit regions 211, which can heat the opposite sides of the preparation unit regions 211, thereby improving the temperature uniformity of the chamber 21 and effectively preventing the sample liquid from freezing due to excessively low temperature during the sample addition process. Of course, the heating electrodes 30 can also be arranged in other ways, which are not limited here.

[0178] It should be noted that the heating electrode 30 can be configured as either a single-layer metal layer or a double-layer metal layer, and this is not limited to either. In this embodiment, we will specifically use a double-layer metal layer heating electrode 30 as an example for further explanation, but this is not a limitation.

[0179] For example, the heating electrode 30 includes a first metal layer and a second metal layer stacked together. The first metal layer is disposed on the side of the support 20 opposite to the substrate 10, and the first metal layer is connected between the second metal layer and the support 20.

[0180] In this application, the heating electrode 30 has a relatively high current cross-sectional density, specifically 5 × 10⁻⁶. 5 A / cm 2 When the heating electrode 30 uses a single-layer metal layer, the material of the single-layer metal layer can be W, Au, Ta, or Cu; when the heating electrode 30 uses a double-layer metal layer, the double-layer metal layer can be a combination of W, Au, Ta, and Cu. Specifically, in some embodiments where the heating electrode 30 uses a double-layer metal layer, the first metal layer is made of tungsten or titanium, and the second metal layer is made of gold. In cryo-electron microscopy, since the sample chip 100 is in a low-temperature environment, in order to achieve rapid heating of the sample liquid and maintain it at a preset temperature, the heating electrode 30 needs to withstand a large heating power. Therefore, tungsten or titanium, which can withstand a large heating power and has good thermal conductivity, is used to make the first metal layer, and gold is used to make the second metal layer.

[0181] Please see Figure 4 For example, a support layer 40 is provided on the side of the substrate 10 opposite to the support member 20. That is, a support layer 40 is provided on the bottom of the substrate 10. In this way, on the one hand, the support layer 40 can improve the structural strength of the sample chip 100; on the other hand, the support layer 40 can ensure the flatness of the bottom surface of the sample chip 100.

[0182] When the sample chip 100 is placed on the grid, the support layer 40 will be in contact with the grid. To improve the thermal stability of the sample chip 100, the support layer 40 in this application is made of a material with a low coefficient of thermal expansion, ensuring the dimensional stability of the sample chip 100 during temperature changes. During transmission electron microscopy observation, the sample needs to be frozen. At this time, the support layer 40 can prevent the sample chip 100 from bending, twisting, or deforming due to temperature changes, avoiding sample displacement or image blurring caused by deformation of the sample chip 100, thus providing a stable observation environment for the experiment.

[0183] Meanwhile, under the action of the support layer 40, the sample chip 100 can be placed flat on the carrier in the transmission electron microscope, ensuring the positional stability of the sample chip 100 in the transmission electron microscope.

[0184] Furthermore, to ensure the propagation of the electron beam, the support layer 40 is provided with a second cutout portion 41. The projection of the first cutout portion 11 along the thickness direction of the support member 20 lies within the outline of the second cutout portion 41. After acquiring information from the sample, the electron beam first passes through the substrate 10 along the first cutout portion 11, then passes through the support layer 40 along the second cutout portion 41, and finally reaches the equipment in the cryo-electron microscope used to receive the electron beam.

[0185] When the injection port 22 is, for example, located on the side of the support 20 facing the substrate 10 and covered by the substrate 10 along the thickness direction of the support 20, in order to add sample liquid into the injection port 22, please refer to... Figures 15 to 20 The sample chip 100 shown. Figures 15 to 20 The sample chip 100 shown is Figures 4 to 14 The sample chip 100 shown has a similar overall structure and performs essentially the same functions; therefore, the identical parts will not be elaborated upon further. The following section will focus on the differences... Figures 4 to 14 The structure of the sample carrier chip 100 shown will be described in detail. For example, a sample inlet channel 12 is provided on the substrate 10. The sample inlet channel 12 penetrates the substrate 10 along its thickness direction and communicates with the sample inlet aperture 22. Thus, sample liquid can be added into the sample inlet channel 12 and enter the sample inlet aperture 22 through the sample inlet channel 12.

[0186] Please see Figure 15Based on the aforementioned embodiments, when a support layer 40 is provided on the side of the substrate 10 away from the support member 20, in order not to affect the addition of sample liquid into the injection channel 12, an injection port 42 is provided on the support layer 40, and the injection port 42 is connected to the injection channel 12.

[0187] Please see Figure 15 and Figure 16 For example, the substrate 10 also includes flow channels. These flow channels are located between the sample inlet channel 12 and the sample inlet 22. The flow channels comprise multiple stages connected in series according to the direction of liquid flow. The number of stages is not limited to one, two, three, or more; it is not limited here and can be flexibly adjusted and set according to actual needs. Specifically, to facilitate the entry of sample liquid into the chamber 21 through the sample inlet 22, the flow channels are configured as at least two stages, with at least two stages connected in series. Along the direction of sample liquid flow, the first flow channel is connected to the sample inlet channel 12, and the last flow channel is connected to the sample inlet 22. Thus, the sample liquid enters the first flow channel through the sample inlet channel 12, then flows along the series-connected flow channels, and is introduced into the chamber 21 through the last flow channel and the sample inlet 22.

[0188] Based on the aforementioned embodiments, the cross-sectional dimensions of the flow channels at different levels decrease along the direction of sample liquid flow. Thus, as the sample liquid flows along each flow channel, the flow resistance decreases with the reduction in the cross-sectional dimensions of the flow channels, allowing the sample liquid to more easily penetrate into the chamber 21 through the inlet 22.

[0189] For example, a recess is formed on the side of the substrate 10 facing the support 20. Specifically, the recess can be formed by gas etching during the fabrication of the substrate 10. The recess is recessed in a direction away from the support 20, and the recess mates with the support 20 to form a flow channel.

[0190] The number of flow channels is the same as the number of recesses, and they are arranged in a one-to-one correspondence. Since the cross-sectional dimensions of different levels of flow channels differ along the flow direction of the sample liquid, the cross-sectional dimensions of the recesses corresponding to different levels of flow channels also differ. The recesses of different levels are sequentially connected, and their cross-sectional dimensions decrease along the flow direction of the sample liquid. Specifically, the depth of the recesses of different levels decreases along the flow direction of the sample liquid.

[0191] Please see Figure 15 and Figure 16In this embodiment, the flow channels are configured in two stages, namely a first flow channel 13 and a second flow channel 14; correspondingly, the recesses are configured in two stages, namely a first recess and a second recess. The first recess corresponds to the first flow channel 13, and the second recess corresponds to the second flow channel 14. The first flow channel 13 is connected to the sample inlet channel 12, and the first flow channel 13 is also connected to the second flow channel 14, which is connected to the sample inlet port 22. The depth of the first recess is greater than the depth of the second recess. Optionally, the depth of the first recess may include, but is not limited to, 2 μm to 5 μm; the depth of the second recess may include, but is not limited to, 1 μm to 2 μm.

[0192] Based on the aforementioned embodiments, for recesses of the same level, such as a first recess or a second recess, the number of first recesses is, for example, at least two, and they are sequentially connected; similarly, the number of second recesses is, for example, at least two, and they are sequentially connected. The more recesses there are, the longer the flow channel corresponding to each recess will be; conversely, when the number of recesses decreases, the length of the flow channel corresponding to each recess will decrease. The arrangement of the recesses will affect and change the arrangement of the flow channels. During the fabrication of the sample chip 100, each recess can be individually processed on the substrate 10 through gas etching, which facilitates the fabrication of flow channels on the substrate 10.

[0193] Based on the aforementioned embodiments, at least two stages of flow channels are spirally arranged along the outer periphery of the support 20, and the distance between the last flow channel and the center of the support 20 is less than the distance between the first flow channel and the center of the support 20. Specifically, taking two flow channels as an example, the two flow channels are the first flow channel 13 and the second flow channel 14. After the first flow channel 13 is arranged around the outer periphery of the support 20, the first end of the second flow channel 14 is connected to the tail end of the first flow channel 13. The second flow channel 14 starts from the tail end of the first flow channel 13 and continues to be arranged along the outer periphery of the support 20, and the second flow channel 14 is located inside the first flow channel 13. In this way, the sample liquid flows and is guided along at least two stages of flow channels, spirally flows around the outer periphery of the support 20, and finally seeps into the chamber 21 through the sample inlet 22, which makes it easier for the sample liquid to be introduced into the chamber 21 and reduces the residue of the sample liquid.

[0194] Based on the aforementioned embodiments, when the chamber 21 is provided with multiple preparation unit areas 211, the arrangement of the multiple preparation unit areas 211 in the chamber 21 is not limited. Specifically, taking the multiple preparation unit areas 211 arranged in a straight line in a direction perpendicular to the thickness direction of the support member 20 as an example, the flow channel at the end is arranged around the periphery of the multiple preparation unit areas 211, and at least two sample inlets 22 are provided, which are arranged sequentially and at intervals along the flow channel at the end. In this way, the sample liquid can enter the chamber 21 through each sample inlet 22 and penetrate into each preparation unit area 211 from different positions, which facilitates penetration into each preparation unit area 211 of the chamber 21, thereby improving sample preparation efficiency.

[0195] For example, at least two injection ports 22 are provided in a one-to-one correspondence with at least two preparation unit areas 211. Each injection port 22 is aligned with each preparation unit area 211. The sample liquid entering the chamber 21 through the injection port 22 can penetrate into the corresponding preparation unit area 211, so that each preparation unit area 211 can be quickly filled with sample liquid, thereby improving sample preparation efficiency and imaging quality.

[0196] Of course, in some alternative solutions, the injection port 22 may not need to correspond to the preparation unit area 211. The number of injection ports 22 may also be more or less than the number of preparation unit areas 211, which is not limited here and can be flexibly adjusted and set according to actual needs.

[0197] Please refer to the following: Figures 15 to 20 Similar to Figure 14 As illustrated, the chamber 21 is provided with a plurality of partitions 25. Each partition 25 is disposed, for example, between two adjacent preparation unit regions 211. In addition, the support member 20 also includes a first support membrane 23 and a second support membrane 24.

[0198] Please see Figures 15 to 20 For example, the support member 20 further includes an intermediate connecting layer 27. The intermediate connecting layer 27 connects the first support film 23 and the second support film 24. A window 271 is provided in the middle of the intermediate connecting layer 27, and the window 271 penetrates the intermediate connecting layer 27 along its thickness direction. The first support film 23, the second support film 24, and the window 271 enclose a cavity 21. The height of the cavity 21 can be adjusted accordingly by adjusting the thickness of the intermediate connecting layer 27.

[0199] When the area of ​​the window 271 in the intermediate connecting layer 27 is large, the portion of the second support membrane 24 corresponding to the window 271 is not effectively and stably supported, leading to adhesion with the first support membrane 23. Therefore, the support member 20 includes support pillars 26. The support pillars 26 are connected to the first support membrane 23 and the second support membrane 24, providing support for the second support membrane 24, improving its stability, and effectively preventing adhesion. Optionally, multiple support pillars 26 can be provided. Each support pillar 26 may correspond to, for example, each preparation unit region 211, or, for example, each sample inlet 22.

[0200] Please see Figure 20 For example, along the thickness direction of the support member 20, the projections of the end flow channel, injection port 22, partition 25, support column 26, first observation section 231 and second observation section 241 on the intermediate connecting layer 27 are all located within the outline of the window 271.

[0201] In some embodiments, please refer to Figures 21 to 24 , Figure 21 It shows Figure 15 A bottom view of an embodiment of the substrate in the sample carrier chip 100 is shown. Figure 15 As can be seen, the substrate 10 has a first cutout portion 11. Because the opening size of the first cutout portion 11 is relatively large, the structural strength of the substrate 10 is reduced, which will affect the support stability of the sample carrier chip 100 to some extent. Therefore, please refer to the comparison. Figures 22 to 24 , Figure 22 It shows Figure 15 Another embodiment of the substrate in the sample carrier chip 100 is shown in a bottom view structural diagram. Figure 23 It shows Figure 15 A bottom view of another embodiment of the substrate in the sample chip 100 shown. Figure 24 It shows Figure 15 The cross-sectional view of the sample carrier chip 100 at the separating and reinforcing portions is shown. Compared to Figure 21 The first hollow portion 11 shown is configured as a complete hollow area. The first hollow portion 11 of the substrate 10 has at least two hollow areas 111. The number of hollow areas 111 includes, but is not limited to, 2, 3, 5, 10 or more, which can be flexibly adjusted and set according to actual needs. Optionally, multiple hollow areas 111 are arranged sequentially at intervals along the length or width direction of the first hollow portion 11. A reinforcing portion 16 is provided between any two adjacent hollow areas 111. The reinforcing portion 16 is connected to the inner wall of the first hollow portion 11, specifically to the inner walls of opposite sides of the first hollow portion 11 along the width direction, thereby increasing the structural strength of the substrate 10 and improving the support stability.

[0202] Based on the aforementioned embodiment, the reinforcing portion 16 and the separating portion 25 are aligned along the thickness direction of the substrate 10. In other words, the reinforcing portion 16 and the fabrication unit region 211 are misaligned along the thickness direction of the substrate 10. This avoids the reinforcing portion 16 from obstructing the image and affecting the observation effect of the sample layer.

[0203] Understandably, not limited to Figure 15 A reinforcing portion 16 may be provided within the first cutout portion 11 of the substrate in the sample carrier chip 100 shown, and for example, a reinforcing portion 16 may be provided within the first cutout portion 11 of the substrate. Figures 4 to 14 In any of the embodiments shown, a reinforcing portion 16 is provided in the first cutout portion 11 of the substrate of the sample chip 100 to increase the structural strength of the substrate 10.

[0204] In some embodiments, please refer to Figure 25 and Figure 26 Compared to Figure 17 and Figure 20 The difference in the structure shown is that the first support membrane 23 is provided with a drainage groove 232. One end of the drainage groove 232 is connected to the sample inlet 22, and the other end extends to the preparation unit area 211. Specifically, there are multiple drainage grooves 232, each corresponding to a sample inlet 22. Under the drainage effect of the drainage grooves 232, the sample liquid at the sample inlet 22 is concentratedly drained to the preparation unit area 211.

[0205] Based on the aforementioned embodiments, in order to reduce the wetting resistance of the sample liquid, the cross-sectional dimensions of the drainage channel 232 decrease along the direction close to the preparation unit region 211. Specifically, as... Figure 25 and Figure 26 As shown, the distance between the two opposite walls of the drainage groove 232 decreases along the direction close to the preparation unit region 211.

[0206] In the description of this application, it should be understood that if terms such as "center", "longitudinal", "lateral", "length", "width", "thickness", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", "clockwise", "counterclockwise", "axial", "radial", "circumferential" appear, these terms indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings, and 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.

[0207] Furthermore, where the terms "first" and "second" appear, these terms are for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined with "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this application, where the term "multiple" appears, "multiple" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0208] In this application, unless otherwise expressly specified and limited, the terms "installation," "connection," "joining," and "fixing," etc., should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral part; 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; they can refer to the internal communication of two components or the interaction between two components, unless otherwise expressly limited. Those skilled in the art can understand the specific meaning of the above terms in this application based on the specific circumstances.

[0209] In this application, unless otherwise expressly specified and limited, the use of descriptions such as "above" or "below" the second feature indicates that the first and second features are in direct contact or indirect contact via an intermediate medium. Furthermore, "above," "on top of," and "over" the second feature can mean that the first feature is directly above or diagonally above the second feature, or simply that the first feature is at a higher horizontal level than the second feature. Similarly, "below," "below," and "under" the second feature can mean that the first feature is directly below or diagonally below the second feature, or simply that the first feature is at a lower horizontal level than the second feature.

[0210] It should be noted that if an element is referred to as being "fixed to" or "set on" another element, it can be directly on the other element or there may be an intervening element. If an element is considered to be "connected to" another element, it can be directly connected to the other element or there may be an intervening element. If so, the terms "vertical," "horizontal," "upper," "lower," "left," "right," and similar expressions used in this application are for illustrative purposes only and do not represent the only possible implementation.

[0211] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0212] The embodiments described above are merely illustrative of several implementation methods of this application, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent application. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this application, and these all fall within the protection scope of this application. Therefore, the protection scope of this patent application should be determined by the appended claims.

Claims

1. A sample preparation apparatus, characterized in that, include: Cooling mechanism; A low-temperature heat sink is used to support the sample carrier unit. A heat conduction mechanism is provided, which is connected between the cooling mechanism and the low-temperature heat sink. The cooling mechanism cools the sample carrier unit supported on the low-temperature heat sink through the heat conduction mechanism and the low-temperature heat sink.

2. The sample preparation apparatus according to claim 1, characterized in that, The heat conduction mechanism includes a cooling braided strip, which is connected between the cooling mechanism and the low-temperature heat sink.

3. The sample preparation apparatus according to claim 2, characterized in that, The cooling braided tape is made of at least one material, including oxygen-free copper or graphite sheets.

4. The sample preparation apparatus according to claim 1, characterized in that, The cooling mechanism includes a liquid gas container.

5. The sample preparation apparatus according to claim 4, characterized in that, The liquid gas container is provided with a liquid storage cavity, the top of which is open so that the liquid gas inside the liquid storage cavity is exposed through the top of the liquid storage cavity.

6. The sample preparation apparatus according to claim 5, characterized in that, The sample preparation device also includes a support frame, which is arranged adjacent to the liquid gas container, and the cryogenic heat sink is installed on the top of the support frame.

7. The sample preparation apparatus according to claim 6, characterized in that, The support frame includes legs and a heat insulation base. The legs are connected to the heat insulation base, and the low-temperature heat sink is installed on the heat insulation base.

8. A sample preparation system, characterized in that, The sample preparation apparatus includes the sample preparation device as described in any one of claims 1 to 7, and further includes a sample carrying unit; the sample carrying unit includes: A heating mechanism for supporting a sample carrier chip and heating the sample carrier chip; A support platform, the support platform being used to support the heating mechanism; and A sample carrier chip includes a substrate and a support member. The support member is connected to the substrate and has a chamber and a sample inlet. The inner wall of the chamber is hydrophilic, and the sample inlet is connected to the chamber.

9. The sample preparation system according to claim 8, characterized in that, The heating mechanism is provided with a positioning groove for positioning the sample chip, and the sample chip is placed in the positioning groove and bonded and fixed to the heating mechanism.

10. The sample preparation system according to claim 8, characterized in that, At least a portion of the support platform is disposed on the low-temperature heat sink, and another portion of the support platform extends out of the low-temperature heat sink and is suspended in the air. The projection of the sample chip along the thickness direction of the support platform is offset from that of the low-temperature heat sink.

11. The sample preparation system according to claim 10, characterized in that, The sample preparation device also includes a positioning frame, a part of which is connected to the low-temperature heat sink, and the other part of which extends out of the low-temperature heat sink and is suspended in the air. The positioning frame is used to support and position the support platform.

12. The sample preparation system according to claim 8, characterized in that, The sample carrying unit further includes a clamping member, which is connected to the carrying platform and is used to press and fix the sample chip and the heating mechanism onto the carrying platform.

13. The sample preparation system according to any one of claims 8 to 12, characterized in that, The heating mechanism includes a heating chip; the sample carrying unit further includes an adapter plate and an electrical connection connector plate; the heating chip is electrically connected to the adapter plate, and the heating chip is stacked on the adapter plate; the adapter plate is disposed on the carrying platform; the adapter plate is electrically connected to the electrical connection connector plate, and the electrical connection connector plate is used for electrical connection with the controller.

14. The sample preparation system according to claim 13, characterized in that, The adapter plate is encapsulated and fixed to the support platform.

15. The sample preparation system according to claim 13, characterized in that, The heating chip is electrically connected and thermally contacted to the metal leads of the adapter board by bonding.

16. The sample preparation system according to claim 8, characterized in that, The outer wall of the support is made of hydrophobic material.

17. The sample preparation system according to claim 8, characterized in that, The chamber is provided with multiple preparation unit areas, and the spacing between the two inner walls of each preparation unit area along the thickness direction of the support is either different or the same.

18. The sample preparation system according to claim 17, characterized in that, The support includes a first support film and a second support film that are spaced apart from each other. The first support film is connected to the substrate. The second support film is located on the side of the first support film away from the substrate and cooperates with the first support film to form the cavity. The first support film is provided with a first observation portion, and the second support film is provided with a second observation portion. The second observation portion is arranged opposite to the first observation portion.

19. The sample preparation system according to claim 18, characterized in that, The substrate has a first cutout portion, and the projections of the first observation portion and the second observation portion along the thickness direction of the support member are both located within the outline range of the first cutout portion.

20. The sample preparation system according to claim 19, characterized in that, The first hollow portion has at least two hollow areas, and a reinforcing portion is provided between any two adjacent hollow areas, the reinforcing portion being connected to the inner wall of the first hollow portion.

21. The sample preparation system according to claim 18, characterized in that, There are multiple first observation sections and multiple second observation sections; each first observation section is correspondingly arranged with each of the preparation unit regions, and each second observation section is correspondingly arranged with each of the preparation unit regions.

22. The sample preparation system according to claim 18, characterized in that, Both the first observation section and the second observation section are elongated, and the width of both the first observation section and the second observation section is set to 0.6μm-2.5μm.

23. The sample preparation system according to claim 18, characterized in that, The first observation section includes a first hole penetrating the first support membrane, and the second observation section includes a second hole penetrating the second support membrane.

24. The sample preparation system according to claim 23, characterized in that, The first support membrane includes a first main support portion and a first auxiliary support portion, the first auxiliary support portion being connected to the first main support portion, and the first hole being disposed in the first auxiliary support portion; The thickness of the first auxiliary support part is less than the thickness of the first main support part; And / or, The second support membrane includes a second main support portion and a second auxiliary support portion, the second auxiliary support portion being connected to the second main support portion, and the second hole being disposed in the second auxiliary support portion; The thickness of the second auxiliary support part is less than the thickness of the second main support part.

25. The sample preparation system according to claim 24, characterized in that, The first main support portion is arranged circumferentially around the first auxiliary support portion; and / or, the second main support portion is arranged circumferentially around the second auxiliary support portion.

26. The sample preparation system according to claim 24, characterized in that, The first observation unit further includes a first sealing film connected to the first support film, the first sealing film being used to seal the first hole, and the first sealing film being transparent under electron beam irradiation; and / or, the second observation unit further includes a second sealing film connected to the second support film, the second sealing film being used to seal the second hole, and the second sealing film being transparent under electron beam irradiation.

27. The sample preparation system according to claim 26, characterized in that, The first sealing film comprises graphene material; and / or, the second sealing film comprises graphene material.

28. The sample preparation system according to claim 18, characterized in that, The support also includes a support column, which is connected between the first support membrane and the second support membrane.

29. The sample preparation system according to claim 28, characterized in that, The support column is integrally formed with at least one of the first support membrane and the second support membrane.

30. The sample preparation system according to claim 8, characterized in that, The substrate has a support layer on the side opposite to the support member.

31. The sample preparation system according to claim 18, characterized in that, The substrate is provided with a sample inlet channel, which penetrates the substrate along the thickness direction and is connected to the sample inlet hole.

32. The sample preparation system according to claim 31, characterized in that, The substrate is further provided with a flow channel, which is located between the sample inlet channel and the sample inlet hole; the flow channel is provided in at least two stages and is arranged in series; along the sample liquid flow direction, the first flow channel is connected to the sample inlet channel, and the last flow channel is connected to the sample inlet hole.

33. The sample preparation system according to claim 32, characterized in that, Along the direction of sample liquid flow, the cross-sectional dimensions of the flow channels at different levels tend to decrease.

34. The sample preparation system according to claim 32, characterized in that, The substrate has a recess on the side facing the support member, the recess being recessed in a direction away from the support member, and the recess cooperating with the support member to form the flow channel.

35. The sample preparation system according to claim 8, characterized in that, It also includes ohmic contact resistors; the ohmic contact resistors are electrically connected to at least one of the substrate and the support; the ohmic contact resistors are configured as two, the two ohmic contact resistors are arranged at intervals, and the two ohmic contact resistors are electrically connected to the positive and negative terminals of the external conductive components, respectively.

36. A method for preparing a sample using the sample preparation system as described in any one of claims 8 to 35, characterized in that, include: The heating mechanism is activated to heat the sample carrier chip, and the temperature of the sample carrier chip is controlled to be higher than the first preset temperature. The sample liquid is added into the chamber through the inlet port, and the sample liquid spontaneously wets the chamber through capillary action. Once the sample liquid wets the entire chamber, the heating mechanism stops working, and the cooling mechanism lowers the temperature of the sample carrier chip to below the second preset temperature, causing the sample liquid inside the sample carrier chip to freeze rapidly.

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