CN113528343B - An adherent cell culturer for terahertz wave irradiation - Google Patents

Abstract

The invention discloses an adherent cell culture vessel for terahertz wave irradiation, comprising a cylindrical chamber for accommodating cell culture fluid and culturing cells, and the bottom wall of the cylindrical chamber can be used for terahertz wave focused beam to pass through. However, the environment for culturing cells is a cell growth microenvironment of 37°C, 5% CO2, and saturated humidity. A convex part is arranged in the cylindrical chamber, and the bottom surface of the convex part is round and fixed with the bottom wall of the cylindrical chamber. Coinciding and sealingly connected, the bottom wall of the cylindrical chamber located between the side wall of the convex part and the inner wall of the cylindrical chamber serves as the annular adherent growth surface of the cells. The invention mainly solves the problem of poor consistency of the biological effect of the terahertz wave of cells with a simple-structured adherent cell culturer.

Description

An adherent cell culturer for terahertz wave irradiation

technical field

The invention belongs to the technical field of terahertz wave biological effects, in particular to an adherent cell culture vessel used for terahertz wave irradiation.

Background technique

In recent years, the application of terahertz technology has become more and more extensive, and the application research of its imaging and sensing technology in medical, military, and security fields has continued to deepen. In order to apply terahertz technology safely and effectively, especially in the fields of diagnosis, in vivo imaging, treatment, security inspection, etc., it is necessary to study and evaluate the biological effects and safety of terahertz radiation, and to clarify its biological effects and characteristics. The dose-response relationship was obtained to obtain the injury threshold. Therefore, the research on the biological effects of terahertz waves has become a hot spot in this field.

In the prior art, document CN111239377A discloses a system and method for long-term in vitro terahertz wave biological effect research, which can realize long-term in vitro irradiation of various primary cells and cell lines that grow adherently. Hertzian waves provide a stable experimental platform and method for studying the biological effects of terahertz waves. The system and method are used to ensure the growth environment of cells when irradiated outside the body only from a constant temperature sample chamber, that is, the temperature, humidity, and carbon dioxide concentration are consistent with the incubator environment, thereby eliminating the possible effects of changes in the external environment on the growth state of cells. However, studies have found that the factors affecting the consistency of cellular effects under terahertz radiation are not only the growth environment, and the system and method still have the problem of poor consistency of the biological effects of cellular terahertz waves.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide an adherent cell culture vessel for terahertz wave irradiation, which is at least used to solve the problem of poor consistency of the biological effects of terahertz waves on cells.

The purpose of the present invention is achieved by adopting the following technical solutions.

An adherent cell culturer for terahertz wave irradiation, comprising a columnar chamber for accommodating cell culture fluid and culturing cells, the bottom wall of the columnar chamber being capable of passing through a focused beam of terahertz waves, and culturing cells The environment is a cell growth microenvironment of 37°C, 5% CO2, and saturated humidity. It is characterized in that: a convex part is arranged in the cylindrical chamber, and the bottom surface of the convex part is round and fixed with the bottom wall of the cylindrical chamber. Coinciding and sealingly connected, the bottom wall of the cylindrical chamber located between the side wall of the convex portion and the inner wall of the cylindrical chamber serves as the annular adherent growth surface of the cells.

In order to improve the stability of the terahertz wave irradiation process, the bottom wall of the cylindrical chamber adopts a rigid polystyrene plastic sheet with a thickness of not more than 0.5 mm after surface modification treatment.

In order to further improve the consistency of the biological effects of terahertz waves, the diameter of the bulge is controlled so that the cell adherent growth surface is completely projected within the radial cross-section range where the strong irradiation is located, that is, the width H of the cell adherent growth surface is smaller than that of the strong irradiation. The terahertz focused beam is generated by a dual-color field laser air plasma terahertz wave irradiation system and focused by necessary optical instruments. The necessary optical instruments include but are not limited to off-axis parabolic mirrors, THz total reflection lenses, and THz lenses.

As a preferred solution, the energy value of the strong irradiation is greater than 1/2 of the maximum energy value of the focused beam of the terahertz wave.

For the convenience of operation, the adherent cell culture apparatus includes a cylindrical shell, the cylindrical chamber is axially arranged on the shell, and the shell is used for engaging in the through hole of the orifice plate, and the hole is The plate is fixed on the irradiation support; the protrusions are cylindrical or conical.

Further, the adherent cell culture vessel includes a cylindrical shell, the cylindrical chamber is axially arranged on the shell, and the shell is directly fixed on the irradiation support; the protruding part is integrally formed. , and the cylindrical part and the conical part are arranged coaxially.

As a preferred embodiment of the present invention, a through hole is provided on the casing, an arc-shaped annular groove is arranged on the inner side wall of the casing near the bottom of the through hole; The circular plate, an elastic annular sealing element is fixed on the edge of the circular plate, the elastic annular sealing element can be embedded in the arc-shaped annular groove, and is jointly enclosed by the circular plate, the side wall of the tapered portion and the inner side wall of the casing. The space for containing the cell culture fluid, and the surface of the circular plate is the bottom wall of the cylindrical chamber. Adopting such a structure can also improve the utilization rate of the casing, which is beneficial to saving consumables. In the present invention, the elastic annular sealing elements are all made of materials that meet medical requirements.

As another preferred embodiment of the present invention, the adherent cell culture vessel comprises a cylindrical shell, the cylindrical chamber is axially arranged on the shell, and the shell is directly fixed on the irradiation support; The protruding portion is cylindrical, and a support lug is arranged on the top surface of the protruding portion; a through hole is arranged on the casing, and an arc-shaped ring groove is arranged on the inner side wall of the casing near the bottom of the through hole; A circular plate suitable for cell adherent growth is arranged at the bottom of the raised portion, and an elastic annular sealing element is fixed on the edge of the circular plate. The elastic annular sealing element can be embedded in the arc-shaped annular groove, and the The side wall of the rising part and the inner side wall of the shell together form a space for containing the cell culture liquid, and the surface of the circular plate is the bottom wall of the cylindrical chamber. Adopting such a structure not only facilitates the auxiliary operations of cell culture, such as disassembling and assembling the circular plate for cell adherent growth, but also facilitates the observation of the growth condition of irradiated cells after terahertz irradiation.

In order to further improve the stability of the terahertz wave irradiation process, the bottom wall of the cylindrical chamber adopts a rigid polystyrene plastic film with a thickness of not more than 0.2 mm and a horizontal plane after surface modification treatment. The protruding portion adopts a sac-shaped body that is filled with liquid and is fully expanded; the connection method between the bottom wall of the cylindrical chamber and the bottom surface of the protruding portion is sealing and bonding.

In order to further improve the stability during the terahertz wave irradiation process, the protrusions and the shell are made of hydrophobic polystyrene materials, which can prevent cells from sticking to the side of the protrusions. wall growth, reducing the error in the consistent evaluation of the biological effects of terahertz waves on irradiated cells.

Beneficial effects: the invention mainly solves the problems of uniformity of irradiated cells and poor consistency of terahertz wave biological effects mainly with a simple-structured adherent cell culturer. The specific structure of the device avoids the terahertz spectrum/light wave energy that does not meet the energy requirements, thereby greatly improving the consistency of the terahertz wave biological effect of the irradiated cells; The cell culture apparatus is beneficial to improve the uniformity and stability of the terahertz wave irradiation process, as well as the convenience of operation, and those of ordinary skill in the art can perform the operation smoothly; For the adherent cell culture vessel of the same batch, the observation can be repeated multiple times and at intervals without changing the cell culture vessel, which is beneficial to improve the accuracy and efficiency of the experimental results.

Description of drawings

1 is a schematic cross-sectional view of an adherent cell culture vessel used for terahertz wave irradiation in Example 1;

2 is a schematic diagram of a disc and a raised portion of an adherent cell culture vessel used for terahertz wave irradiation in Example 1;

3 is a schematic cross-sectional view of an adherent cell culture vessel used for terahertz wave irradiation in Example 2;

4 is a schematic diagram of a disc and a raised portion of an adherent cell culture vessel used for terahertz wave irradiation in Example 2;

5 is a schematic diagram of an adherent cell culture vessel used for terahertz wave irradiation in Example 3;

Fig. 6 is the schematic diagram of the orifice plate in embodiment 3;

7 is a schematic diagram of an adherent cell culture vessel used for terahertz wave irradiation in Example 4;

8 is a schematic diagram of an adherent cell culture vessel used for terahertz wave irradiation in Example 5;

Fig. 9 is the partial schematic diagram of terahertz wave irradiation optical path in embodiment 1-5;

10 is a schematic diagram of a light spot after focusing of the terahertz wave irradiation optical path in Embodiments 1-5;

H in Fig. 1 represents the width H of the cell adherent growth surface, and h in Fig. 9 represents the width/diameter of the weakly irradiated region.

Detailed ways

The present invention will be further described below in conjunction with specific embodiments. It is pointed out that the following embodiments should not be construed as limiting the protection scope of the present invention. Those of ordinary skill in the art make some non-essential improvements and adjustments according to the content of the present invention, which are all in the present invention. within the scope of protection.

Example 1

1 and 2, an adherent cell culture vessel for terahertz wave irradiation includes a cylindrical shell 7, which is used to be directly fixed on the irradiation support; on the shell 7 A cylindrical chamber 2 is axially arranged, and the cylindrical chamber 2 is substantially a part of the through hole on the housing 7. A cylindrical convex portion 3 is provided in the cylindrical chamber 2, and the convex portion 3 is arranged in the cylindrical chamber 2. The top surface of the part 3 is provided with a support lug 33, which is mainly used for clamping tools such as clamps or tweezers. The bottom surface of the raised part 3 is circular and is fixed, overlapped and sealed with the bottom wall of the cylindrical chamber 2. The bottom wall of the cylindrical chamber 2 located between the side wall of the raised portion 3 and the inner wall of the cylindrical chamber 2 serves as the annular adherent growth surface of the cells. An arc-shaped annular groove 11 is arranged on the inner side wall of the casing 7 near the bottom of the through hole, a circular plate 10 suitable for cell adherent growth is arranged at the bottom of the raised portion 3 , and an elastic annular sealing element 12 is fixed on the edge of the circular plate 10 , the elastic annular sealing element 12 can be embedded in the arc-shaped annular groove 11, and the space for containing the cell culture liquid is jointly enclosed by the circular plate 10, the side wall of the raised portion 3 and the inner side wall of the housing 7, and the surface of the circular plate 10 is cylindrical The bottom wall of the chamber 2, the bottom wall of the cylindrical chamber 2 (that is, the circular plate 10) can be passed through by the terahertz focused beam, that is, the terahertz focused beam can pass through the circular plate 10 and irradiate on the adherent cells to produce biological effect.

Among them, the bottom wall (circular plate 10 ) of the cylindrical chamber 2 adopts a rigid polystyrene plastic sheet with a thickness of not more than 0.3 mm after surface modification treatment, and the rigid polystyrene plastic sheet after surface modification treatment is used. It is conducive to the growth of cells adherent to the wall. The raised part 3 and the shell 7 are made of hydrophobic polystyrene material. Adherent growth.

The principle of irradiating adherent cells with terahertz waves in this embodiment: the terahertz wave irradiation optical path 21 generated by a two-color field laser air plasma terahertz irradiation system and focused by necessary optical instruments is distributed in a conical shape, such as As shown in FIGS. 9 and 10 , along the radial section of the cone, the photon energy near the axis 26 is weak, and this part of the photon energy is difficult to meet the irradiation requirements for adherent cells, and the peak number 25 in FIG. 9 refers to The trough represents the energy of the focused beam of the terahertz wave, and the peak area 22 (that is, the strong irradiation area, the energy value of this area is greater than 1/2 of the maximum value of the energy of the focused beam of the terahertz wave), the energy of the terahertz spectrum/light wave is strong, The valley area 23 (that is, the weak irradiation area, the energy value of this area is less than 1/2 of the maximum energy value of the terahertz wave focused beam) The energy of the terahertz spectrum/light wave is weak, and the conical top is the terahertz wave irradiation light path In the focused light spot 24 , the energy in the middle area of the light spot 24 is weaker corresponding to the wave valley area 23 , and the energy in the area outside the middle of the light spot 24 is stronger and corresponds to the wave peak area 22 . In this embodiment, a raised portion 3 is provided on the circular plate 10 , so that cells can only adhere to the bottom wall of the cylindrical chamber 2 between the side wall of the raised portion 3 and the inner wall of the cylindrical chamber 2 . , and by controlling the diameter of the raised portion 3, the cell adherent growth surface is completely projected within the radial cross-section range where the strong irradiation is located, that is, the width H of the cell adherent growth surface is smaller than the width of the radial cross-section where the strong irradiation is located, The area where the cells adhere to the wall just corresponds to the peak area 22 with strong energy, so as to cleverly avoid the terahertz spectrum/light wave energy that does not meet the energy requirements, and greatly improve the uniformity of the cells irradiated and the terahertz wave biological effect consistency.

Before the terahertz wave irradiation is performed, the circular plate 10 with the convex part 3 is clamped in the arc-shaped annular groove 11 in the cylindrical chamber 2, and the environment for culturing the cells adopts a temperature of 37°C, 5% CO2, and saturated humidity. Cell growth microenvironment, after the cells in the cylindrical chamber 2 have adhered to the appropriate stage, transfer them to a microscope for observation or take samples for observation under a glass slide (usually, a non-polluting environment is required) Observation below), after the observation is over, it will continue to the next stage of cultivation. Since the edge of the circular plate 10 is fixedly provided with the elastic annular sealing element 12, when the circular plate 10 needs to be taken out under special circumstances, the elastic annular sealing element 12 is still attached to the circular plate 10 after the circular plate 10 is removed. The cells on the plate 10 provide a space for storing the cell growth solution, which is also conducive to cell growth during the observation stage. For the same batch of adherent cells, the observation can be repeated multiple times and at intervals without changing the cell culture vessel. It is beneficial to improve the accuracy and efficiency of experimental results. Moreover, the process of disassembling and assembling the circular plate 10 is simple and convenient, and those skilled in the art can perform the operation smoothly.

Example 2

An adherent cell culture vessel for terahertz wave irradiation, as shown in Example 1 and in conjunction with Figure 3 and Figure 4, the main difference between it and Example 1 is that the structure of the protruding part 3 is different. Compared with Example 1, the circular plate 10 of the cell culture apparatus is easier to disassemble: the adherent cell culture apparatus includes a cylindrical shell 7, the cylindrical chamber 2 is arranged on the shell 7, and the shell 7 is directly fixed on the radial. The protruding part 3 is composed of a cylindrical part 32 and a conical part 31 which are integrally formed and arranged coaxially. The cylindrical part 32 is easy to hold during the assembly or disassembly process; An arc-shaped annular groove 11 is arranged on the inner side wall of the casing 7 near the bottom of the through hole; a circular plate 10 suitable for cell adherent growth is arranged at the bottom of the tapered portion 31, and an elastic annular seal is fixed on the edge of the circular plate 10. Element 12, the elastic annular sealing element 12 can be embedded in the arc-shaped annular groove 11, and is jointly enclosed by the circular plate 10, the side wall of the tapered portion 31 and the inner side wall of the shell 7 to form a space for holding the cell culture liquid. The surface of the circular plate 10 is: Cylindrical chamber 2 bottom wall.

Example 3

An adherent cell culture vessel for terahertz wave irradiation, as shown in Figures 5 and 6, includes a housing 7, the housing 7 is provided with a lug 5 that is convenient to handle, and the housing 7 is used for clamping. It is combined in the through hole 8 of the orifice plate 9, the orifice plate 9 is fixed on the irradiation bracket, and is arranged on the shell 7 for accommodating the cell culture medium and culturing the cells. The cylindrical chamber 2, the bottom wall of the cylindrical chamber 2 It can pass through the terahertz focused beam, and the environment for culturing cells is a cell growth microenvironment of 37°C, 5% CO2, and saturated humidity; a convex part 3 is arranged in the cylindrical chamber 2, and the convex part 3 is cylindrical , the bottom surface of the raised portion 3 is round and is fixed, overlapped and sealed with the bottom wall of the cylindrical chamber 2, and the bottom wall of the cylindrical chamber 2 is located between the side wall of the raised portion 3 and the inner wall of the cylindrical chamber 2 As the annular adherent growth surface of the cells.

Among them, the bottom wall of the cylindrical chamber 2 is a rigid polystyrene plastic sheet with a thickness of not more than 0.3 mm after surface modification treatment, and the rigid polystyrene plastic sheet after surface modification treatment is conducive to cell adhesion. The protruding part 3 and the shell 7 are made of hydrophobic polystyrene material, and the hydrophobic polystyrene material can prevent the cells from adhering to the side wall of the protruding part 3 and the shell 7 from growing.

The principle of irradiating adherent cells with terahertz waves in this example is similar to that in Example 1. Other advantages of the adherent cell culture apparatus in this embodiment mainly include: easy access to a single component 6 with a cell culture medium and a raised portion 3, and the orifice plate 9 can be reused, which is beneficial to saving the orifice plate.

Example 4

An adherent cell culture vessel for terahertz wave irradiation, as shown in Example 3 and in conjunction with FIG. 7 , the main difference between it and Example 1 is that the structure of the raised part 3 is different: the raised part 3 is in the shape of a cone shape, and the apex of the convex part is lower than the top of the shell 7 .

Example 5

An adherent cell culture vessel for terahertz wave irradiation, as shown in Example 3 and in conjunction with FIG. 8 , the main difference from Example 1 is that the structure of the raised part 3 is different: the raised part 3 is in the shape of a cone shape, and the apex of the raised portion is flush with the top of the housing 7 .

In addition, a cover is also provided on the top of the housing 7, and the cover is well covered during the cell culture process to prevent the environment in the cylindrical chamber 2 from being polluted.

Claims (7)

1. The utility model provides an adherent cell culture ware for terahertz wave irradiation, is including being used for holding cell culture solution and cultivateing cylindrical cavity (2) of cell, and cylindrical cavity (2) diapire can supply terahertz wave focusing beam to pass, cultivates the cell environment be 37 ℃, 5% CO2, the cell growth microenvironment of saturation humidity, its characterized in that: a bulge part (3) is arranged in the cylindrical chamber (2), the bottom surface of the bulge part (3) is circular and is fixed with the bottom wall of the cylindrical chamber (2), overlapped and hermetically connected, and the bottom wall of the cylindrical chamber (2) between the side wall of the bulge part (3) and the inner wall of the cylindrical chamber (2) is used as an annular adherent growth surface of cells; the bottom wall of the cylindrical chamber (2) is a horizontal hard polystyrene plastic thin plate with the thickness not more than 0.3mm after surface modification treatment, or the bottom wall of the cylindrical chamber (2) is a horizontal hard polystyrene plastic thin film with the thickness not more than 0.2mm after surface modification treatment; the diameter of the bulge part (3) is controlled to enable the annular adherent growth surface of the cell to be completely projected in the radial section range where strong irradiation is located, a terahertz wave irradiation light path (21) which is generated by a double-color field laser air plasma terahertz irradiation system and focused by a necessary optical instrument is adopted to be distributed in a conical shape, and the energy value of the strong irradiation is larger than 1/2 of the energy maximum value of the terahertz wave focused beam.

2. The adherent cell culture device for terahertz wave irradiation of claim 1, wherein: the adherent cell culture device comprises a cylindrical shell (7), the cylindrical chamber (2) is axially arranged on the shell (7), the shell (7) is clamped in a through hole (8) of a pore plate (9), and the pore plate (9) is fixed on an irradiation support; the convex part (3) is cylindrical or conical.

3. The adherent cell culture device for terahertz wave irradiation of claim 1, wherein: the adherent cell culture device comprises a cylindrical shell (7), the cylindrical chamber (2) is axially arranged on the shell (7), and the shell (7) is directly fixed on the irradiation support; the boss (3) is composed of a cylindrical portion (32) and a conical portion (31) which are integrally formed and coaxially arranged.

4. The adherent cell culture device for terahertz wave irradiation of claim 3, wherein: a through hole is formed in the shell (7), and an arc-shaped ring groove (11) is formed in the inner side wall of the shell (7) close to the bottom of the through hole; conical part (31) bottom is provided with plectane (10) that are fit for the cell adherent growth, is provided with elasticity annular sealing element (12) at plectane (10) border fixed, and this elasticity annular sealing element (12) can imbed in arc annular (11), by plectane (10) conical part (31) lateral wall with casing (7) inside wall encloses synthetic splendid attire cell culture liquid's space jointly, plectane (10) surface does cylindricality cavity (2) diapire.

5. The adherent cell culture device for terahertz wave irradiation of claim 4, wherein: the adherent cell culture device comprises a cylindrical shell (7), the cylindrical chamber (2) is axially arranged on the shell (7), and the shell (7) is directly fixed on the irradiation support; the boss (3) is cylindrical, and a support lug (33) is arranged on the top surface of the boss (3); a through hole is formed in the shell (7), and an arc-shaped ring groove (11) is formed in the inner side wall of the shell (7) close to the bottom of the through hole; bellying (3) bottom is provided with plectane (10) that are fit for the cell adherent growth, is provided with elasticity annular sealing element (12) at plectane (10) border fixed, and this elasticity annular sealing element (12) can imbed in arc annular (11), by plectane (10) bellying (3) lateral wall with casing (7) inside wall encloses the space of synthetic splendid attire cell culture liquid jointly, plectane (10) surface does cylindricality chamber (2) diapire.

6. The adherent cell culture device for terahertz wave irradiation of claim 5, wherein: the bulge (3) adopts a sac body which is full of liquid and is inflated fully; the bottom wall of the cylindrical cavity (2) is connected with the bottom surface of the bulge (3) in a sealing bonding mode.

7. The adherent cell culture device for terahertz wave irradiation of any one of claims 5 to 6, wherein: the bulge (3) and the shell (7) are both made of hydrophobic polystyrene materials.

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