Biological material supercooling storage system and working method thereof

Abstract

The invention discloses a biological material supercooling storage system and a working method thereof. The biological material supercooling storage system comprises a sample carrier for storing and placing a biological material; the sample cavity is used for providing a subcooled storage environment of-30 to-5 DEG C for the biological material; the temperature control module is used for providing a controllable cooling rate for the interior of the refrigeration module, the cooling rate is greater than 2 DEG C / min in a temperature interval of above 5 DEG C, the cooling rate is 1-2 DEG C / min in a temperature interval of 0-5 DEG C, and the cooling rate is less than 1 DEG C / min in a temperature interval of below 0 DEG C; the electromagnetic excitation module is used for generating a low-frequency to very high-frequency 50Hz-300MHz electromagnetic field in the sample, and inhibiting the formation and growth of ice crystals in the sample by using the non-thermal effect of the electromagnetic field; the cooperative control module is used for achieving cooperative control over the cooling rate, the supercooling temperature and the electromagnetic field intensity so as to maintain long-term storage of the samples in a liquid state at the supercooling temperature, the storage temperature of the biological materials is reduced, meanwhile, the biological materials are not frozen, and liquid-state storage of the biological materials with different sources and different sizes at the low temperature can be achieved.

Description

Technical Field

[0001] This invention relates to the field of biomedical engineering technology, and in particular to a supercooled preservation system for biomaterials and its working method. Background Technology

[0002] Cell therapy, represented by stem cells and immune cells, has achieved good results in modern medicine, especially in the treatment of immune system diseases, hematological diseases, and malignant tumors. Transplantation of organs such as the heart and liver is the ultimate solution for many organ failure conditions. Furthermore, the demand for biological cells is very strong in fields such as biomedical research, genetic information expression, and drug testing and development. However, these biomaterials need to be obtained from living tissues or organs, which are not only scarce but also rapidly degenerate and become inactive after being removed from the body, resulting in waste. Lowering the storage temperature of cells is an effective way to maintain cell viability; for every 10°C decrease in temperature, the metabolic rate is reduced by half. Currently, cryopreservation is still the main method of low-temperature biotechnology, which inevitably produces ice crystals during the cooling process, thus damaging cell morphology and function. At the same time, the rewarming process can also cause physical damage such as decrystallization and uneven thermal stress. Therefore, supercold preservation technology, which lowers the storage temperature of biological materials without causing them to freeze, will be an effective method to extend the storage period of biological materials. Summary of the Invention

[0003] This invention aims to at least partially solve one of the technical problems in related technologies. Therefore, one objective of this invention is to provide a supercooled preservation system for biological materials and its operating method, which lowers the preservation temperature of biological materials without causing them to freeze, meeting the differentiated temperature requirements of different biological materials and providing a comprehensive preservation solution for cells, tissues, organs, and even samples from different sources.

[0004] In a first aspect, the present invention proposes a supercooled preservation system for biomaterials, comprising a sample carrier for storing and placing samples, a sample chamber, a temperature control module, an electromagnetic excitation module, and a collaborative control module;

[0005] The sample chamber is used to provide a supercooled preservation environment of -30 ~ -5℃ for biological materials;

[0006] The temperature control module is used to provide a controllable cooling rate for the sample chamber. The cooling rate is greater than 2℃ / min in the temperature range above 5℃, 1~2℃ / min in the temperature range of 0~5℃, and less than 1℃ / min in the temperature range below 0℃, and finally balanced at the preset supercooled storage temperature.

[0007] The electromagnetic excitation module is placed inside the sample cavity and is used to generate a low-frequency to very high-frequency electromagnetic field (50Hz ~ 300MHz) within the sample, with an electric field strength of 0 ~ 10.5 V / m, magnetic induction intensity 0 ~ 5mT, and utilize the non-thermal effect of electromagnetic field to suppress the formation and growth of ice crystals in the sample;

[0008] The collaborative control module is used to achieve collaborative control of cooling rate, supercooling temperature and electromagnetic field strength, so as to maintain the sample in liquid state for a long time at supercooling temperature.

[0009] Preferably, the sample carrier includes a microfluidic device for carrying discrete phase samples and a bag or tube device for carrying continuous phase samples.

[0010] Preferably, the sample carrier is a microfluidic device, and the electromagnetic excitation module uses a flat electrode; the sample carrier is a bag-type or tubular device, and the electromagnetic excitation module uses an electromagnetic coil.

[0011] Preferably, a door is installed at the opening of the sample cavity, and multiple sets of electromagnetic excitation modules are provided inside the sample cavity. The sample carrier is placed inside the magnetic field generated by the electromagnetic excitation module. A through hole is opened in the sample cavity corresponding to the wire connection of the electromagnetic excitation module. A signal source is externally connected to the electromagnetic excitation module by a wire passing through the through hole. A platform is provided at the upper end of the electromagnetic excitation module. A placement port is opened on the platform corresponding to the position of the sample carrier. The sample carrier is detachably connected to the placement port.

[0012] Preferably, the sample cavity has an air inlet for connecting to an external cooling duct and an air outlet for discharging gas on its side wall. The temperature control mechanism includes a cooling duct and a heat exchanger for cooling the cooling duct. The air outlet of the cooling duct is connected to the air inlet of the sample cavity, and a fan is installed at the air inlet of the cooling duct.

[0013] Preferably, the collaborative control module has a control console inside, and the power adjustment switches for the fan, heat exchanger and signal source are all located at the control console.

[0014] Preferably, the platform is equipped with a temperature sensor and a Hall sensor for detecting the internal temperature of the sample carrier and the magnetic field strength near the sample carrier.

[0015] Secondly, the present invention provides a method for operating a supercold preservation system for biological materials, comprising any of the above-mentioned schemes for supercold preservation systems for biological materials, wherein the method steps are as follows:

[0016] S1: Add the biological materials and the required culture medium or protectant into the sample carrier, and seal it to prevent microbial infection;

[0017] S2: Open the sample chamber door, place the sample carrier at the designated position on the stage, and close the door;

[0018] S3: Start the heat exchanger and fan in sequence via the control console. After the air temperature is reduced by cooling water, the air is sent to the sample chamber through the air inlet. After cooling the sample chamber, the air is sent out through the air outlet.

[0019] S4: Start the signal source via the control console to provide an excitation signal of a preset frequency to the electromagnetic excitation module;

[0020] S5: During the preservation process, the control console uses temperature sensors and Hall sensors to detect the temperature and magnetic field strength of the sample carrier in real time, and coordinates the output power of the heat exchanger, fan and signal source to maintain the sample in a liquid state for a long time at a supercooled temperature.

[0021] The beneficial effects of this invention are:

[0022] (i) This invention achieves supercooled preservation of biological materials, that is, the solution does not freeze after reaching its freezing point, thereby reducing ice crystal damage to biological materials and extending the preservation time of biological materials.

[0023] (ii) The present invention proposes an electromagnetic field-assisted supercooling preservation method, which improves the thermodynamic stability of existing supercooling preservation technology.

[0024] (iii) The supercooling preservation method for biological materials described in this invention can achieve supercooling preservation of biological materials of different volumes. Attached Figure Description

[0025] In the attached diagram:

[0026] Figure 1 This is a schematic diagram of the structure of a biomaterial supercold preservation system based on an electromagnetic coil proposed in this invention;

[0027] Figure 2 This is a schematic diagram of the structure of a biomaterial supercold preservation system based on a flat plate electrode proposed in this invention;

[0028] Figure 3 This is a schematic diagram of the sample cavity structure proposed in this invention;

[0029] Figure 4 This is an example diagram of the cooling rate control proposed in this invention;

[0030] Figure 5 This is an example diagram of electromagnetic field strength control proposed in this invention;

[0031] Figure 6 This is an example diagram illustrating the suppression of ice crystal formation by alternating electromagnetic fields at -10℃ proposed in this invention.

[0032] In the diagram: 1-fan, 2-heat exchanger, 3-refrigeration module, 4-electromagnetic coil, 5-sample carrier, 6-signal source, 7-control console, 8-stage, 9-air inlet, 10-air outlet, 11-flat electrode. Detailed Implementation

[0033] Reference Figure 1 , Figure 2 and Figure 3 A supercooled preservation system for biomaterials includes a sample carrier 5 for storing and placing samples, a sample chamber 3, a temperature control module, an electromagnetic excitation module, and a collaborative control module.

[0034] Sample chamber 3 is used to provide a cold preservation environment for biological materials;

[0035] Specifically, the cold storage environment temperature should be in the range of -30 ~ -5℃.

[0036] The temperature control module is used to provide a controllable cooling rate for the interior of the refrigeration module 3;

[0037] Specifically, the cooling rate is greater than 2℃ / min in the temperature range above 5℃, 1~2℃ / min in the temperature range of 0~5℃, and less than 1℃ / min in the temperature range below 0℃, ultimately balancing at the preset supercooled storage temperature.

[0038] The electromagnetic excitation module is located inside the cooling module 3 of the temperature control module. It is used to generate low-frequency to very high-frequency electromagnetic fields in the sample under the excitation of the signal source, and to use the non-thermal effect of the electromagnetic field to suppress the formation and growth of ice crystals in the sample.

[0039] Specifically, the electromagnetic field frequency is 50Hz ~ 300MHz, and the electric field strength is 0 ~ 10. 5 V / m, magnetic induction intensity 0 ~ 5mT.

[0040] The collaborative control module is used to achieve coordinated control of cooling rate, supercooling temperature and electromagnetic field strength to maintain the sample in a liquid state for a long time at supercooling temperature.

[0041] In this embodiment: the sample carrier 5 includes a microfluidic device for carrying discrete phase samples and a bag or tube device for carrying continuous phase samples;

[0042] Specifically, discrete phase samples can be microdroplets, while continuous phase samples can be large-scale solutions.

[0043] In this embodiment: the sample carrier 5 is a microfluidic device, and the electromagnetic excitation module adopts a flat electrode 11; the sample carrier 5 is a bag-type or tube-type device, and the electromagnetic excitation module adopts an electromagnetic coil 4.

[0044] Specifically, microfluidic devices can be specific materials with special functional requirements, such as microfluidic chips; bag or tube devices can be general and readily available experimental consumables such as test tubes, glass bottles, and storage bags.

[0045] In this embodiment: a door is installed at the opening of the sample cavity 3, multiple electromagnetic excitation modules are provided inside the sample cavity 3, the sample carrier 5 is placed inside the magnetic field generated by the electromagnetic excitation module, a through hole is opened at the connection of the electromagnetic excitation module wire in the sample cavity 3, a signal source 6 is connected to the electromagnetic excitation module by a wire passing through the through hole, a platform 8 is provided at the upper end of the electromagnetic excitation module, a placement port is opened at the position of the platform 8 corresponding to the sample carrier 5, and the sample carrier 5 is detachably connected to the placement port.

[0046] The electromagnetic excitation module provides a uniformly distributed AC electromagnetic field for the supercooled metastable state. It utilizes the non-thermal effect of the electromagnetic field to enhance molecular perturbation, thereby inhibiting ice crystal growth. The electromagnetic excitation module can be an electromagnetic generator such as an electromagnetic coil 4, a flat plate electrode 11, or a Helmholtz coil.

[0047] The electromagnetic excitation module provides an electromagnetic field to assist in supercooling storage, utilizing the polarization effect of the dielectric to suppress ice crystal formation. Its polarization intensity can be expressed as... ;

[0048] in, ε0 is the polarizability, which describes the polarization capability under the action of an electric field and is related to the frequency; ε0 is the vacuum permittivity; and E is the applied electric field strength.

[0049] In this embodiment: the sample cavity 3 has an air inlet 9 for connecting to the external cooling air duct and an air outlet 10 for discharging gas on its side wall. The temperature control mechanism includes a cooling air duct and a heat exchanger 2 for cooling the cooling air duct. The air outlet of the cooling air duct is connected to the air inlet 9 of the sample cavity 3. A fan 1 is installed at the air inlet of the cooling air duct.

[0050] Specifically, the cooling air is directly obtained from the environment by the fan 1. The air volume of the fan 1 is adjustable. The heat exchanger 2 can adjust the cooling efficiency of the cooling duct by adjusting the power of the cooling water pump, so as to control the cooling rate of the sample chamber 3.

[0051] In this embodiment: the air inlet 9 of the sample cavity 3 is located on the lower part of one side of the sample cavity 3, and the air outlet 10 of the sample cavity 3 is located on the upper part of the other side of the sample cavity 3.

[0052] Specifically, the air inlet 9 is located at the bottom, and the air outlet 10 is located at the top. The cooling airflow can cool the sample carrier 5 from bottom to top, thus improving the cooling effect.

[0053] In this embodiment: the collaborative control module is equipped with a control console 7, and the power adjustment switches of the fan 1, heat exchanger 2 and signal source 6 are all located at the control console 7.

[0054] Specifically, the power adjustment switches for fan 1, heat exchanger 2, and signal source 6 are all located on control panel 7 for easy operation. Simultaneously, by controlling the flow rate of fan 1 and water pump, the requirements of the biological materials in the sample chamber for different supercooling temperatures (-30 ~ -5℃) and cooling rates are met. The current of signal source 6 is controlled to meet the requirements of the biological materials in the sample chamber for different electromagnetic fields (50Hz ~ 300MHz, 0 ~ 10). 5 The requirements of V / m and 0 ~ 5mT, and the coordinated control of the temperature field in the electromagnetic field.

[0055] In this embodiment: a temperature sensor and a Hall sensor for detecting the internal temperature and magnetic field strength of the sample carrier 5 are provided at the position of the stage 8 near the sample carrier 5.

[0056] Specifically, temperature and magnetic field strength information are fed back in real time by temperature sensors and Hall sensors to regulate the fan 1, water pump, and signal source 6.

[0057] As another embodiment of this application, this embodiment proposes a working method for a biological material supercold preservation system, which includes any of the above-mentioned solutions for biological material supercold preservation systems.

[0058] Reference Figure 1 , Figure 1 Electromagnetic coil 4 is used as the electromagnetic excitation module, suitable for bag or tubular devices carrying continuous phase (large-scale solution) samples. The working method is as follows:

[0059] S1: Add the biological material and the required culture medium or protectant into the sample carrier 5, and seal it to prevent microbial infection;

[0060] S2: Connect the electromagnetic coil 4 to the pre-reserved hole in the sample cavity, place the loaded sample carrier 5 in the middle of the stage 8, open the sample cavity door, place the stage 8 inside the sample cavity, and close the door.

[0061] S3: Through the control console 7, heat exchanger 2 and fan 1 are started in sequence. After the air temperature is reduced by cooling water, it is sent to the sample chamber through the air inlet 9. After cooling the sample chamber, it is sent out through the air outlet 10. Fan 1 draws air from the environment and exchanges heat with cooling water through heat exchanger 2, thereby generating cold air of -30 ~ -5℃ to provide to the sample chamber.

[0062] S4: Through the control console 7, the signal source 6 is activated to provide an excitation signal of a certain frequency to the electromagnetic coil 4, and further, to provide an electromagnetic field of a certain intensity to maintain the stability of the supercooled preservation of biological materials.

[0063] S5: The control console 7 adjusts the flow rate of the heat exchanger 2 and the fan 1 by controlling the subcooling of the outer wall of the sample carrier 5 and the air, thereby controlling the cooling rate.

[0064] by Figure 4 Taking the illustrated embodiment as an example, under the conditions of an air temperature of -30℃ and a flow rate of 0.37kg / h, the cooling rate of the sample carrier 5 is 5℃ / min from 20 to 5℃, 1℃ / min from 5 to 0℃, and 0.02℃ / min from 0 to -10℃.

[0065] S6: The console 7 adjusts the excitation signal of the signal source 6 by reading the Hall sensor, thereby controlling the electromagnetic field strength.

[0066] like Figure 5 As shown, with the excitation current frequency being 1MHz, the left figure shows an excitation current of 1A, and the right figure shows an excitation current of 2A. The maximum magnetic induction intensity in the cooling cavity is 0.6mT in the left figure and 0.6mT in the right figure. Figure 1 0.2mT.

[0067] In this embodiment, as Figure 2 As shown, the other method steps are the same as those in the above embodiments, except that: the microfluidic device suitable for carrying discrete phase (microdroplet) samples uses a flat plate electrode 11 as an electromagnetic excitation module.

[0068] like Figure 6 As shown, the effect diagram of electromagnetic-assisted supercooling preservation is given. The device was continuously cooled for 1.5 hours in a low temperature environment of -10℃ (left figure), and no ice formed under the condition of electromagnetic field (50Hz, 10V) (right figure).

Claims

1. A supercold preservation system for biological materials, characterized in that: It includes a sample carrier (5) for storing and placing samples, a sample chamber (3), a temperature control module, an electromagnetic excitation module, and a collaborative control module; The sample chamber (3) is used to provide a supercooled preservation environment of -30 ~ -5℃ for biological materials; The temperature control module is used to provide a controllable cooling rate for the sample cavity (3). The cooling rate is greater than 2℃ / min in the temperature range above 5℃, 1~2℃ / min in the temperature range of 0~5℃, and less than 1℃ / min in the temperature range below 0℃, and finally balanced at the preset supercooled storage temperature. The electromagnetic excitation module is placed inside the sample cavity (3) and is used to generate a low-frequency to very high-frequency electromagnetic field of 50Hz to 300MHz within the sample, with an electric field strength of 0 to 10. 5 V / m, magnetic induction intensity 0 ~ 5mT, and utilize the non-thermal effect of electromagnetic field to suppress the formation and growth of ice crystals in the sample; The collaborative control module is used to achieve collaborative control of cooling rate, supercooling temperature and electromagnetic field strength, so as to maintain the sample in liquid state for a long time at supercooling temperature.

2. The supercold preservation system for biological materials according to claim 1, characterized in that: The sample carrier (5) includes a microfluidic device for carrying discrete phase samples and a bag or tube device for carrying continuous phase samples.

3. The biomaterial supercold preservation system according to claim 2, characterized in that: The sample carrier (5) is a microfluidic device, and the electromagnetic excitation module adopts a flat electrode (11); the sample carrier (5) is a bag-type or tube-type device, and the electromagnetic excitation module adopts an electromagnetic coil (4).

4. The biomaterial supercold preservation system according to claim 1, characterized in that: The sample cavity (3) is equipped with a door at the opening position. Multiple sets of electromagnetic excitation modules are provided inside the sample cavity (3). The sample carrier (5) is located inside the magnetic field generated by the electromagnetic excitation module. The sample cavity (3) is provided with a through hole at the connection point of the electromagnetic excitation module wire. The electromagnetic excitation module is connected to a signal source (6) by a wire passing through the through hole. The upper end of the electromagnetic excitation module is provided with a platform (8). The platform (8) is provided with a placement port at the position of the sample carrier (5). The sample carrier (5) is detachably connected to the placement port.

5. A biomaterial supercold preservation system according to claim 4, characterized in that: The sample cavity (3) has an air inlet (9) for connecting to an external cooling duct and an air outlet (10) for discharging gas on its side wall. The temperature control mechanism includes a cooling duct and a heat exchanger (2) for cooling the cooling duct. The air outlet of the cooling duct is connected to the air inlet (9) of the sample cavity (3). A fan (1) is installed at the air inlet of the cooling duct.

6. The supercold preservation system for biomaterials according to claim 3, characterized in that: The collaborative control module is equipped with a control console (7), and the power adjustment switches of the fan (1), heat exchanger (2) and signal source (6) are all located at the control console (7).

7. A biomaterial supercold preservation system according to claim 5, characterized in that: The platform (8) is equipped with a temperature sensor and a Hall sensor for detecting the internal temperature and magnetic field strength of the sample carrier (5) near the sample carrier (5).

8. A method for operating a supercold preservation system for biological materials, characterized in that: The method comprises a biomaterial supercold preservation system as described in any one of claims 1-7, wherein the steps are as follows: S1: Add the biological material and the required culture medium or protectant into the sample carrier (5), and seal it to prevent microbial infection; S2: Open the door of the sample chamber (3), place the sample carrier (5) at the designated position on the platform (8), and close the door; S3: Through the control console (7), start the heat exchanger (2) and the fan (1) in sequence. After the air temperature is reduced by cooling water, it is sent to the sample chamber (3) through the air inlet (9). After cooling the sample chamber (3), it is sent out through the air outlet (10). S4: Start the signal source (6) through the console (7) to provide an excitation signal of preset frequency to the electromagnetic excitation module; S5: During the preservation process, the control console (7) detects the temperature and magnetic field strength of the sample carrier (5) in real time through temperature sensor and Hall sensor, and coordinates the output power of heat exchanger (2), fan (1) and signal source (6) to maintain the sample in liquid state for a long time at supercooled temperature.

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