阅读说明：本技术 一种高效的超大量液槽池制备方法 (Efficient preparation method of ultra-large liquid tank ) 是由 陈斌 刘志文 于 2021-08-13 设计创作，主要内容包括：本发明公开了一种高效的超大量液槽池制备方法,以碳酸二甲酯(DMC)为溶剂制备浓度为15%-20%(w/v,g/mL)的聚醋酸乙烯酯(PVAC)溶液作为壳前驱液,以水为溶剂制备10 vol%-70 vol%甘油溶液作为核前驱液,通过同轴静电纺丝技术,以碳支持膜直接接收纺丝纤维,从而得到由PVAC包裹水的纤维状可直接用于电子显微镜测试的液槽池。本方法可以在30 s内制备约9000个液槽池,极大地提高了液槽池的制备效率,整个流程中所使用的原材料价格低廉易获取,大幅度节约了成本；同时制备的大量液槽池都在同一个支撑膜或基底上,因而可在同样的外界条件下观测大量样品在真实液体环境下的行为,从而可以方便的用来做统计研究。(The invention discloses a high-efficiency preparation method of an ultra-large liquid tank, which is characterized in that a polyvinyl acetate (PVAC) solution with the concentration of 15-20% (w/v, g/mL) is prepared by taking dimethyl carbonate (DMC) as a solvent to serve as a shell precursor solution, a glycerol solution with the concentration of 10-70 vol% is prepared by taking water as a solvent to serve as a core precursor solution, and a carbon supporting film is used for directly receiving spinning fibers through a coaxial electrostatic spinning technology, so that the fibrous liquid tank which is wrapped by the PVAC and can be directly used for electron microscope testing is obtained. The method can prepare about 9000 liquid tank pools within 30 s, greatly improves the preparation efficiency of the liquid tank pools, has low price and easy acquisition of raw materials used in the whole process, and greatly saves the cost; meanwhile, a large number of prepared liquid tank pools are on the same supporting film or substrate, so that the behavior of a large number of samples in a real liquid environment can be observed under the same external condition, and the method can be conveniently used for statistical research.)

1. An efficient preparation method of an ultra-large liquid tank is characterized by comprising the following steps: taking 15-20% (w/v, g/mL) of polyvinyl acetate solution as a shell precursor solution, and taking water as a solvent to prepare 10-70 vol% of glycerol solution as a core precursor solution; and spinning on the support membrane by an electrostatic spinning technology after the core precursor solution and the shell precursor solution are prepared.

2. The method for preparing ultra-large number of liquid tanks according to claim 1, wherein the step of electrospinning is performed by an electrospinning machine, after the core precursor liquid and the shell precursor liquid are prepared, the core precursor liquid and the shell precursor liquid are respectively transferred to corresponding injectors of a coaxial electrospinning machine, the temperature is kept at 30-40 ℃ under the standard pressure, the relative humidity is 30-40%, the core precursor liquid and the shell precursor liquid are made to form a taylor cone through the coaxial electrospinning machine and generate the final core-shell nano fiber, the voltage during spinning is 10-20 kV, the distance from the needle point of a coaxial needle to a receiver is 10-20 cm, and a carbon supporting film is adopted to directly receive the spun fiber, thereby obtaining the fibrous liquid tank coated with water by polyvinyl acetate.

3. The method of claim 1, wherein the shell precursor solution is dimethyl carbonate as a solvent.

4. The method for preparing ultra-large number of fluid cells according to claim 1, wherein the method for preparing the fluid cell sample containing nanoparticles comprises: corresponding nanoparticles are doped in the core precursor solution.

5. Use of a cell prepared by the method of any one of claims 1 to 4 for real-time observation by an electron microscope.

Technical Field

The invention belongs to the technical field of electron microscope detection, and particularly relates to a liquid tank which can be directly used for electron microscope testing.

Background

The electron microscope can carry out real-time observation on the atomic level on the structure and evolution dynamics of materials and biological samples. Since the electron microscope chamber requires a vacuum environment, the samples typically used for characterization are solid-state. However, many real service conditions are in liquid environment, such as electrochemical processes, catalysis, self-assembly, etc. of material systems and various physiological processes of biological systems. The liquid tank is adopted to observe the sample behavior under the liquid by an electron microscope. The bath cells are generally composed of two sheets, between which a liquid sample is injected and sealed. The difficulty of the preparation of the liquid tank pool is as follows: 1) its imaging window must be thin enough to allow the electron beam to pass through; 2) at the same time, such thin imaging window materials must be strong enough to withstand the pressure differential associated with high vacuum. For decades of electron microscopy invention, researchers have since first attempted to seal liquids with thin aluminum foils in the shape of a sandwich, and have been developing to now use silicon nitride/silicon and graphene to make the bath cells.

The more important developments in recent years are summarized below. In 2003, William et al have first reported that using silicon nitride/silicon as a bath window ensures stability under high vacuum with high strength, while lower atomic number also improves spatial resolution in liquid environments. In 2009, Zheng et al produced independent baths with thinner silicon nitride films with resolution of up to 1 nm. In the same year, De Jonge et al added a microtube and a syringe pump to the silicon nitride bath so that the liquid inside the bath could be circulated, thereby successfully observing the state of a large number of samples in the bath. In 2012, Yuk et al prepared a liquid bath by wrapping liquid with double-layer graphene to observe the growth of platinum, and the excellent resolution was attributed to the ultra-thin graphene material. In addition, there are also some graphene-derived baths, such as laying graphene directly on an amorphous carbon film.

Although the development of fluid cells in one direction, such as resolution, is continuously improving, the preparation method of these fluid cells is generally very tedious, time-consuming, labor-consuming and costly, resulting in limited use of the fluid cells. In particular, when a large number of sample cells are required to be able to perform statistical behavior studies, the above means for making the cell cells will be insufficient. Therefore, a simplified preparation method of the liquid bath pool is urgently needed, so that sample statistics under a liquid environment is possible.

Disclosure of Invention

Aiming at the defects in the prior art, the invention develops a novel liquid bath pool preparation technology based on a coaxial electrostatic spinning technology so as to achieve the purposes of efficient and ultra-large-scale preparation, low cost and ready-to-use.

The preparation method of the liquid tank pool comprises the following steps:

(1) raw materials: polyvinyl acetate (170000 g/mol, Macklin), dimethyl carbonate (99%, Adamas), glycerol (99% or more great), ultrapure water.

(2) The production process flow chart is shown in FIG. 4;

(3) the production process comprises the following steps:

1. the liquid tank pool is prepared by adopting a coaxial electrostatic spinning technology, and core-shell precursor liquid needs to be prepared respectively before spinning. Preparing a polyvinyl acetate (PVAC) solution with the concentration of 15-20% (w/v, g/mL) by taking dimethyl carbonate (DMC) as a solvent to serve as a shell precursor solution, and preparing a glycerol solution with the concentration of 10-70 vol% by taking water as a solvent to serve as a core precursor solution (for preparing a liquid tank sample containing nanoparticles, only corresponding nanoparticles need to be doped into the core precursor solution). After preparing the core-shell precursor solution, respectively transferring the solution to corresponding injectors of an electrostatic spinning machine for a subsequent spinning process;

2. the core-shell injector is placed on a micro pump capable of accurately controlling the flow rate, and the temperature is kept at 30-40 ℃ under the standard pressure, and the relative humidity is 30-40%. The core solution and the shell solution are formed into a taylor cone by a coaxial electrospinning technique and the final core-shell nanofibers are produced. The voltage during spinning is 10-20 kV, and the distance from the needle point of the coaxial needle to the receiver is 10-20 cm;

3. the spinning fibers were directly received with a carbon support membrane, resulting in a fibrous bath of water encapsulated by PVAC. The liquid bath pool prepared by the method can be directly used for electron microscope testing.

The technical scheme of the invention has the following advantages:

1. the invention can prepare about 9000 liquid bath pools in 30 s, thus greatly improving the preparation efficiency of the liquid bath pools;

2. the raw materials used in the whole process are low in price and easy to obtain, so that the cost is greatly saved;

3. the prepared large amount of liquid bath pools are all on the same supporting membrane, so that the behavior of a large amount of samples in the liquid bath pools can be observed under the same condition, and the liquid bath pools can be conveniently used for statistical research.

Description of the drawings:

FIG. 1 is a transmission electron microscope viewing image of the liquid bath prepared by the invention under different states;

FIG. 2 is a schematic diagram of in-situ observation of changes in bubbles in a liquid bath according to the present invention;

FIG. 3 is a schematic diagram of the variation of the fluid bath cell according to the invention when containing nanoparticles;

FIG. 4 is a flow chart of the production process of the present invention.

Detailed Description

1. Dispersing 4 g of PVAC in 20 mL of DMC to prepare 20% (w/v) dispersion as a shell solution; dissolving 1 mL of glycerol in 9 mL of ultrapure water to prepare a solution with the concentration of 10 vol% as a nuclear solution;

2. the heater and dehumidifier were turned on and the temperature and relative humidity were controlled at 35 ℃ and 35%, respectively. A20 nm thick piece of carbon support film was taken and its edge was attached to a roller receiver. Adjusting the distance from the needle point of the coaxial needle to the receiver to be 15 cm;

3. two 10 mL syringes were used to take enough core solution and shell solution, respectively, and placed on a micro pump, and the syringes were connected to the core/shell inlet of a coaxial needle, respectively. Firstly, the core/shell solution is pushed forwards at a higher speed to enable the core solution to reach a needle opening, and the overflowing core/shell solution is wiped off;

4. the material pushing speed of the core solution is preset to be 1.12 mL/h, the material pushing speed of the shell solution is preset to be 4.38 mL/h, the spinning voltage is 15 kV, and the rotation speed of the roller is 60 r/min. And sequentially starting the micro pump and the voltage, and in the electrohydrodynamic process of electrostatic spinning, the liquid forms Taylor coning at the needle mouth to form jet flow. The shell solution solvent will quickly volatilize to form a polymer shell, and the core solution will be retained in the fibrous shell to form a bath pool;

5. after spraying for 30 s, the voltage, the micropump and the roller are sequentially turned off, and the carbon support film is taken down for electron microscope testing.

To confirm the working effect of the bath, we placed it on a transmission electron microscope (TEM, TALOS F200X) for characterization. By using an electron beam in an electron microscope as an excitation source, we can observe the dynamic behavior of a sample in a liquid bath tank after being excited in real time through a Charge Coupled Device (CCD) camera, and examples of several typical applications are as follows. FIG. 1: a typical liquid bath tank in a transmission electron microscope. (a) Multiple sink pool images. (b) Magnified images of individual bath cells. (c) The structural schematic diagram of the liquid tank pool. (d) The preparation of a plurality of liquid tank pools on the same substrate realizes the simultaneous observation of the evolution process of a plurality of sample systems (such as bubbles). Here three bubbles under electron beam excitation are demonstrated. These bubbles contract at different rates, with the rates of change being bubble 1 > bubble 3 > bubble 2 in sequence. The length of each bubble is in nanometers.

FIG. 1a shows a typical image of a multi-well, with several linear fibers visible, wherein the wells are located within protruding beads (marked with solid arrows). The image of the cell is further enlarged, and the bubbles inside can be observed as shown in FIG. 1b (the dark part marked by the solid arrow is liquid, and the light part marked by the dotted arrow is bubbles). According to the shape of the bubbles, the prepared liquid tank can be judged to be tubular. The structure of the liquid bath pool is schematically shown in figure 1c, namely a tubular liquid channel wrapped by a PVAC polymer shell.

Compared with the prior liquid bath pool preparation method in the literature, the method can efficiently prepare a large number of liquid bath pools on the same substrate, and the number of the liquid bath pools on one carbon support membrane with the diameter of 3 mm is about 9000 (the spinning time is 30 s). This also allows the dynamic course of the samples in multiple bath cells to be observed at the same time and under the same external conditions, as shown in FIG. 1 d. In the initial state (0 s), three separate baths each contain a bubble whose length is 1950, 1325, 2700 nm, respectively. After 8 seconds of electron beam irradiation, the size of the bubble changed, wherein the bubble 2 and the bubble 3 contracted about 6% and 66% in length, respectively, and the bubble 1 contracted and disappeared. After 16 s of electron beam irradiation, the bubble 2 still exists and the bubble 3 shrinks again to disappear. This means that these bubbles of different sizes will change at different rates when they resist electron beam irradiation, proving that the method can be used to statistically study the dynamic behaviour of a large number of cuvette samples under the same ambient conditions.

We further examined different bubble kinetic behavior in situ, including processes of bubble growth, bubble contraction, bubble stabilization and bubble fusion (fig. 2). A series of electron microscope snapshots showed the evolution of the bubbles over time. (a) Bubbles larger than the critical size tend to grow under electron beam excitation. (b) Bubbles smaller than the critical size tend to shrink. (c) The appropriate size of the bubble (to reach a stable critical size) can remain stable. (d) When multiple bubbles coexist, they merge into a single bubble, which then gradually shrinks until it eventually disappears. The length of each bubble at different times is in nanometers.

For example, for bubble growth (FIG. 2 a), the initial length of 1466 nm of bubbles gradually increased after electron beam irradiation, and the length increased to 2857 nm (approximately 2 times) after 6 s of irradiation. On the other hand, a certain number of bubbles will decrease in length, such as 2421nm bubbles in fig. 2b, which gradually shrink during the excitation process and finally disappear at 10 s. However, at the right size (stable critical size), the original bubble is stable during irradiation (fig. 2 c). Furthermore, if multiple bubbles are present in the same cuvette cell at the same time (fig. 2 d), they will coalesce and form a single bubble after irradiation (3 s) and then shrink until they finally disappear (12 s).

The above demonstrates the application of one field of the prepared bath, i.e. the study of the various kinetic behavior of the bubbles. Meanwhile, the driving force generated by the evolution of the bubbles can drive various motions of the nanoparticles in the liquid, and the method has potential application prospects in the aspects of autonomous nano motors and drug delivery. For the large bubble here we have chosen two typical systems of similar size, namely single gold nanoparticles (symmetric form;. 50 nm; fig. 3a and 3c) and gold aggregates (asymmetric form;. 70 nm; fig. 3b and 3d), demonstrating the effect of the morphological asymmetry effect on the nanostructure sample motion. Fig. 3 (a, b) are snapshot of the gold nanoparticles and the aggregates moving under the transmission electron microscope, from which the movement traces of the particles and the aggregates can be obtained and then the diffusion behavior thereof can be analyzed. And (c, d) shown in fig. 3 are transmission electron microscope images of single gold nanoparticles and gold aggregates in the liquid bath pool and enlarged images in corresponding red dotted line frames, and gold particles with the size of 10 nm can be clearly distinguished.

The movement of these gold nanostructure samples in the liquid is caused by the liquid flow due to the irradiation of the electron beam, which starts to shrink. Therefore, the motion state of the nano structure in the liquid can be observed in real time through the product.

In summary, the method is suitable for efficiently preparing a large number of liquid tanks and carrying out statistical observation. Compared with a silicon nitride film or a graphene film which is a hundred per thousand yuan, the method greatly saves the material cost. Meanwhile, 9000 liquid tank pools can be obtained within 30 s, and time cost is greatly saved.