CN103674678A - Microchannel chip and device for low-temperature preservation of biomaterial - Google Patents
Microchannel chip and device for low-temperature preservation of biomaterial Download PDFInfo
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Abstract
The invention discloses a microchannel chip and device for low-temperature preservation of a biomaterial. The microchannel chip for low-temperature preservation of the biomaterial comprises two wall bodies and an annular gasket, wherein the two wall bodies clamp the annular gasket to form a closed microcavity; a biomaterial inlet and a biomaterial outlet are formed in one wall body and are communicated with the closed microcavity; a plurality of groove type channels are formed in the outer surface of each wall body and provides channels and guide for flow of a work fluid such as a refrigerant and a rewarming agent, so that circular flow of the work fluid is facilitated, the heat exchange efficiency between the work fluid and the biomaterial is improved, and the cooling or rewarming speed is increased; and meanwhile, the low-temperature preservation of the biomaterial by the microchannel chip adopts closed preservation, and pollution and artificial damage of the biomaterial are avoided. The microchannel chip and device for the low-temperature preservation of the biomaterial are simple in structure and can be produced massively.
Description
Technical field
The invention belongs to field of biomedicine technology, be specifically related to a kind of biomaterial low temperature and preserve micro-channel chip and device.
Background technology
It is the long-term unique effective means of preserving of the biomaterials such as current numerous cell, tissue that low temperature is preserved, and in the various fields of national economy, is all just bringing into play irreplaceable effect.At clinical medicine domain, the long-term low temperature of extensive biomaterial is preserved, and is one of effective way solving cell, tissue-derived problem, is also the basis of numerous modern Clinics; At field of scientific study, by application of cold temperature store method, on the one hand can be for scientific research continues to provide various cell resources, also avoided most possibly on the other hand the too fast inefficacy of cell under normal temperature or refrigerated condition and the wasting of resources that causes; In addition, setting up the low temperature of rare vegeto-animal reproduction cell in imminent danger or seed and preserve storehouse, is also the effective ways that continue species, maintain bio-diversity.
Although biomaterial can be preserved in the lower long-term safety of ultralow temperature (as-196 ℃), in cooling (by normal temperature to ultralow temperature) and rewarming (by ultralow temperature to normal temperature) link, be very likely subject to serious damage, i.e. so-called low temperature injury.About the mechanism of low temperature injury, low temperature is preserved educational circles and be there is no final conclusion, but generally admit the aspects such as eucaryotic cell structure damage that the inside and outside ice-crystal growth of cell causes at present, is the main cause of low temperature injury.
" vitrifacation store method " is a kind of desirable Cryopreservation proposing in recent years.So-called " vitrifacation " is a concept in physics, refer to when water or solution fast cooling to form under extremely low temperature a kind of have full-bodied between liquid and solid-state between amorphous state.By impelling biomaterial that glass transition occurs in temperature-fall period, the low temperature injury that can effectively avoid the factors such as the inside and outside ice-crystal growth of cell to cause.But the glass transition of biomaterial needs cooling, the rewarming rate of superelevation.In temperature-fall period, while only having the critical cooling rate of solution inside and outside rate of temperature fall is greater than cell, could there is glass transition in biomaterial smoothly; In rewarming process, rewarming rate also must just can be avoided vitrified solution generation recrystallization phenomenon higher than critical rewarming rate.
In order to obtain the required superelevation of glass transition, rewarming rate falls, in standard method, use tubule (French mini-Straw, 0.25ml) carrying micro-example direct plunge into Liquid Nitrogen (196 ℃), by the huge temperature difference between sample and liquid nitrogen and liquid nitrogen, the vaporization on tubule surface drives sample to lower the temperature rapidly, in rewarming link, tubule is directly dropped in water bath with thermostatic control (approximately 30 ℃), with the temperature difference between sample and warm water, drive sample to be rapidly heated, on this basis, the people such as Vajta have improved the structure of tubule, open stretched thin-tube method (Open Pulled Straw has been proposed, OPS), resulting rate of temperature fall improves greatly.Reubinnoff has also applied this method and has successfully carried out first in the preservation of stem cell vitrifacation low temperature, follow-up study has been carried out lasting improvement to this method, the methods such as seal type stretched thin-tube method (Closed Pulled Straw, CPS), micro-suction nozzle method (Micropipette), Cryotop and Cryotip have successively been there is.
No matter how structure is improved, and the various containers that carry sample in above method all can be to sample and hydraulic fluid, as the heat exchange between cold-producing medium (liquid nitrogen) or rewarming agent (warm water) plays certain thermal resistance effect in cooling or rewarming process.In order to eliminate the impact of container, in another part research, adopted the mode of direct contact, make micro-example directly contact with cold-producing medium or rewarming agent, realize higher heat exchange efficiency.Typical method bag Electronic Speculum net method (Electron Mmicroscope Grids, EMG), cold around-France (Cryoloop) etc., carry micro-cell with meticulous copper mesh and nylon ring respectively.In recent research, also there is a kind of cell micro drop method (Micro Droplet, MD), utilize micro-fluidic chip to generate the droplet of parcel cell, and directly splash in liquid nitrogen and lower the temperature.These class methods can obtain the higher rewarming rate that falls, but need to solve the recovery of cell and the pollution problem that directly contact brings.
Generally speaking, although vitrifacation Cryopreservation research has obtained a series of achievements, existingly fall, rewarming technology is still immature, show:
I. heat exchange efficiency is limited.Existing cool-down method is all that the pool boiling liquid nitrogen (Pool boiling) obtains higher rate of temperature fall by sample from essence, but in pool boiling process, can produce a large amount of liquid nitrogen steamdrums is rolled in around sample, further cooling is produced to heat insulation effect, so the coefficient of heat transfer is generally less than 103W/m2.K; Concerning rewarming process, existing Thawing Methods is generally driven and is heated up by the temperature difference between sample and water bath with thermostatic control, and heat exchange efficiency is lower, and along with sample temperature gos up, the temperature difference reduces, rewarming rate is decayed rapidly, is easy to produce the problems such as devitrification;
II. restricted to sample volume.In classic method, sample is hemisphere or right cylinder, increase along with sample volume, the heat transfer distances that arrives sample interior increases sharply, the obtainable maximum rate of temperature fall of institute also reduces greatly, sample interior lowering temperature uniform can not guarantee, therefore existing method generally by reduce sample volume obtain essential fall, rewarming rate, but the volume of sample can not unrestrictedly dwindle, and too small sample volume not only increases running cost and difficulty, also lost Clinical practicability.
III. system has opening, is difficult to avoid the pollution to sample.
Summary of the invention
The object of the invention is to overcome the problems referred to above of the prior art, provide a kind of biomaterial low temperature that can realize fast cooling and rewarming to preserve micro-channel chip and device.
For solving the problems of the technologies described above, the present invention by the following technical solutions:
A kind of biomaterial low temperature is preserved micro-channel chip, comprise two wall bodies and an annular gasket, two wall body clamping annular gaskets form airtight microcavity, wherein a wall body is provided with biomaterial entrance and biomaterial outlet, biomaterial entrance and biomaterial outlet all communicate with airtight microcavity, are equipped with several grooved passages on the outside surface of two wall bodies.
Further, the arrangement of set grooved channel parallel and length and micro-channel chip wall body are equal in length.
Further, described biomaterial entrance and biomaterial outlet lay respectively on two diagonal angles of biomaterial entrance and biomaterial outlet place wall body.
Further, rectangular, semicircle, triangle of the xsect of described grooved passage etc.
Further, described wall body is silicon chip or sheet metal.
Further, the material of described annular gasket is flexible high molecular material.
Further, the material of described annular gasket is dimethyl silicone polymer (polydimethylsiloxane, PDMS).
Further, a kind of biological material low temperature storage device, comprise the described micro-channel chip of preserving for biomaterial low temperature, also comprise airtight lagging casing, described micro-channel chip level is fixed on the central authorities of lagging casing, the lagging casing one side parallel with micro-channel chip is provided with hydraulic fluid entrance, hydraulic fluid outlet, biomaterial entrance and biomaterial outlet, wherein, the position of the position of biomaterial entrance and biomaterial outlet and set biomaterial entrance on micro-channel chip and biomaterial outlet is corresponding and by pipeline, exported and be communicated with the biomaterial entrance arranging on micro-channel chip and biomaterial respectively, hydraulic fluid entrance is corresponding with the position at place, set grooved passage two ends on micro-channel chip respectively with the position of hydraulic fluid outlet.
Further, the middle part of described lagging casing is provided with a dividing plate, the center of dividing plate has the hole that a size is mated with micro-channel chip size, and this hole is used for laying micro-channel chip, exports corresponding position be respectively equipped with engraved structure on dividing plate with hydraulic fluid entrance and hydraulic fluid.
Compared with prior art, the invention has the beneficial effects as follows:
(1) biomaterial low temperature preservation micro-channel chip of the present invention has higher cooling and/or rewarming rate.The correlative study such as electronic radiation, refrigeration shows, the multiple heat hydrodynamic interaction under microscale can produce high heat exchanger efficiency, and relatively existing pool boiling technology has greater advantage; In addition, the sample of preserving exists with form of film in system of the present invention, and spheroid relatively of the prior art or right cylinder form have larger effective heat exchange area under same sample volume, and the Distance Shortened of heat transmission.Comprehensive above two aspects, based on of the present inventionly falling, Thawing Methods is expected to more effectively realize vitrifacation low temperature and preserves that required superelevation is fallen, rewarming rate;
(2) biomaterial low temperature of the present invention is preserved the low temperature preservation that micro-channel chip can be realized neatly different coolings and/or rewarming rate and realize flexibly different volumes sample.In the present invention, heat transmission in cooling and/or rewarming process mainly concentrates on the thickness direction of biomaterial film, and the thickness of liquid film can be by the size Control of chip Intermediate gasket, therefore by selecting the pad of different size, the present invention can realize different coolings and/or rewarming temperature speed flexibly; On the other hand, can't affect the rewarming rate that falls of sample controlling the yardstick changing on the basis of thickness of liquid film on other both directions;
(3) biomaterial low temperature of the present invention is preserved micro-channel chip to the closed preservation of saving as of biomaterial, effectively avoids pollution and the damage of biomaterial;
(4) biomaterial low temperature preservation micro-channel chip of the present invention and biological material low temperature storage device are mainly made based on micro-processing and MEMS technique, be suitable for producing in enormous quantities, and can be integrated with other microfluidic system, effectively improve the automatization level of processing procedure, improve treatment effeciency, and reduce the additional injuries that people's factor causes.
Accompanying drawing explanation
Fig. 1 is the decomposition texture schematic diagram that the biomaterial low temperature in the embodiment of the present invention is preserved micro-channel chip;
Fig. 2 is the perspective view that the biomaterial low temperature in the embodiment of the present invention is preserved micro-channel chip;
Fig. 3 is the perspective view of the biological material low temperature storage device in the embodiment of the present invention;
Fig. 4 is biomaterial cooling and the rewarming simulation result schematic diagram of the biological material low temperature storage device based in the embodiment of the present invention.
Embodiment
In order to make object of the present invention, technical scheme and advantage clearer, below in conjunction with drawings and Examples, the present invention is further elaborated.Should be appreciated that specific embodiment described herein, only in order to explain the present invention, is not intended to limit the present invention.
As shown in Figure 1, biomaterial low temperature in the present embodiment is preserved micro-channel chip and is comprised 11,12 and annular gaskets 13 of wall body, wall body 11 and wall body 12 clamping annular gaskets 13 form airtight microcavity, wall body 11 is provided with biomaterial entrance 111 and biomaterial outlet 112, biomaterial entrance 11 and biomaterial outlet 112 all communicate with airtight microcavity, and the structure after assembling as shown in Figure 2.During work, biomaterial to be preserved enters airtight microcavity from biomaterial entrance 111, under the restriction in airtight microcavity space, biomaterial is present in airtight microcavity with liquid film form, because liquid film has larger specific surface area, therefore, when the biomaterial low temperature preservation micro-channel chip of employing the present embodiment carries out the low temperature preservation of biomaterial, easily obtain higher fast cooling and/or rapid rewarming speed, and, can also be by controlling the height of the airtight microcavity of THICKNESS CONTROL of annular gasket 13, and then the thickness of controlling formed biomaterial liquid film obtains desirable fast cooling and/or rapid rewarming speed.On the outside surface of wall body 11 and wall body 12, being equipped with several is arranged in parallel and length and wall body grooved passage 14 equal in length, arranging of grooved passage 14 reduces the thermal resistance effect of wall body in cooling and/or rewarming process on the one hand, be on the other hand, for working fluid provides runner as cooling fluid or rewarming fluid, to realize circulation temperature lowering or the rewarming of hydraulic fluid.Take temperature-fall period as example, during cooling, by cooling fluid, as liquid nitrogen is injected into one end of grooved passage 14, the liquid nitrogen that enters grooved passage 14 is the other end downstream to grooved passage 14 in the guide effect of grooved passage 14, in this course, on micro-channel chip surface, there is forced convection and boiling vaporization in liquid nitrogen, and between the biomaterial in micro-channel chip, carry out rapidly heat interchange, and liquid nitrogen and liquid nitrogen steam that the heat of the biomaterial of exchange is flowed are taken away rapidly, so move in circles, realize the rate of temperature fall of superelevation.
In order to guarantee the smooth injection of biomaterial, above-mentioned biomaterial entrance 111 and biomaterial outlet 112 are preferably disposed on its place wall body, on two of wall body 11 diagonal angles.
For the ease of processing, the shape of the xsect of the grooved passage 14 in the present embodiment is preferably rectangle, can certainly be semicircle, other shapes such as triangle.
In order to reduce costs to greatest extent, the wall body 11,12 in the present embodiment is preferably common silicon chip or sheet metal, and the material of annular gasket 13 is preferably common flexible high molecular material dimethyl silicone polymer.
In order better to realize the guide effect of 14 pairs of hydraulic fluids of grooved passage, the present embodiment also provides a kind of concrete biological material low temperature storage device of preserving micro-channel chip based on above-mentioned biomaterial low temperature, as shown in Figure 3, this biological material low temperature storage device comprises biomaterial low temperature preservation micro-channel chip 1 and airtight lagging casing 2, the central authorities that micro-channel chip 1 level is fixed on lagging casing 2, lagging casing 2 one side 21 parallel with micro-channel chip 1 is provided with hydraulic fluid entrance 22, hydraulic fluid outlet 23, biomaterial entrance 24 and biomaterial outlet 25, wherein, the position of biomaterial entrance 24 and biomaterial outlet 25 is corresponding with the position of set biomaterial entrance 111 on micro-channel chip 1 and biomaterial outlet 112, biomaterial entrance 24 is communicated with the biomaterial entrance 111 arranging on micro-channel chip 1 by pipeline, biomaterial outlet 25 exports 112 by pipeline and the biomaterials that arrange on micro-channel chip 1 and is communicated with, and makes in the outer direct injection micro-channel chip 1 of lagging casing 2 of the biological material low temperature storage device that biomaterial can be from the present embodiment.The position of hydraulic fluid entrance 22 and hydraulic fluid outlet 23 corresponds respectively to the position at the place, two ends of 1 set grooved passage 14 on micro-channel chip.
The using method of this biological material low temperature storage device is as follows:
(1) by biomaterial entrance 24, to micro-channel chip 1 is interior, inject biomaterial to be preserved;
(2) after biomaterial is filled with, close biomaterial entrance 24 and biomaterial outlet 25, then, by hydraulic fluid entrance 22 to the interior injection working fluid of lagging casing 2 as liquid nitrogen;
(3) after having lowered the temperature, micro-channel chip 1 is taken out and drop into liquid nitrogen container from biological material low temperature storage device, preserve for a long time;
(4) when needs rewarming, micro-channel chip 1 is retightened in the lagging casing 2 of putting biological material low temperature storage device, by working fluid, if warm water (approximately 40 ℃) is from Working-fluid intaking 22 injections, the single-phase convection heat exchange effect by warm water in micro-logical 14 realizes the rapid rewarming of micro-channel chip 1 interior biomaterial.
For the ease of fixing of micro-channel chip 1, a kind of simple implementation is that a dividing plate 26 is set at the middle part of lagging casing 2, at dividing plate 26 center, offer the hole that a size is mated with micro-channel chip 1 size, be used for laying and fixing micro-channel chip 1, as shown in Figure 3, simultaneously, for the ease of hydraulic fluid, flow through the grooved passage of micro-channel chip 1 lower surface, on dividing plate 26, also have two engraved structures 261 and 262, engraved structure 261 is corresponding with the position of hydraulic fluid entrance 22 and hydraulic fluid outlet 23 with 262 position; Another kind of implementation can arrange by the bottom at lagging casing 2 several supports and realize micro-channel chip 1 fixing in lagging casing 2.The fixed form that it is emphasized that micro-channel chip 1 is not limited to above-mentioned two kinds of implementations, and other structures with identical function all fall into protection scope of the present invention.
The composition of general biomaterial is mainly aqueous solution.After adding cryoprotective agent, the thermophysical property of biomaterial is close to " water+salt+cryoprotective agent " ternary solution.Further hypothesis cryoprotective agent used is EG, and concentration is 45%, according to known solution thermophysical property, can calculate cooling and the rewarming process in prediction biomaterial biological material low temperature storage device in the present embodiment by theory.If the height of the airtight microcavity of micro-channel chip 1 is 200 microns, prediction acquired results as shown in Figure 4.
From result, in temperature-fall period, biological sample to be preserved can pass through dangerous temperature region (Danger Temperature Range, DTR) in 0.042 second, and the temperature province of crystallization most possibly occurs, and average rate of temperature fall reaches 1.0 * 10
5more than ℃/min; In rewarming process, biological sample to be preserved can pass through dangerous temperature region in 0.057 second, and average rewarming rate reaches 0.74 * 10
5℃/min.According to disclosed documents and materials, the rate of temperature fall that currently used standard microtubule method can reach is about 2500 ℃/min, and rewarming rate is about 1300 ℃/min.Contrast shows, the present invention can realize the rewarming rate that falls of superelevation, and prior art has larger performance advantage relatively, and the vitrifacation low temperature that is therefore expected to more effectively to realize biomaterial is preserved.
In addition, the preservable biological material specimens volume of biological material low temperature storage device in the present embodiment can reach a milliliter magnitude, if micro-channel chip 1 length and width are 1cm * 1cm, conservation object volume is about 0～0.2ml, if micro-channel chip 1 length and width are 2cm * 2cm, conservation object volume is about 0～0.8ml, and prior art conservation object is only microlitre or the upgrading of receiving, so the biological material low temperature storage device in the present embodiment also has greater advantage aspect biomaterial volume to be preserved.
Those of ordinary skill in the art will appreciate that, embodiment described here is in order to help reader understanding's principle of the present invention, should be understood to that protection scope of the present invention is not limited to such special statement and embodiment.Those of ordinary skill in the art can make various other various concrete distortion and combinations that do not depart from essence of the present invention according to these technology enlightenments disclosed by the invention, and these distortion and combination are still in protection scope of the present invention.
Claims (9)
1. a biomaterial low temperature is preserved micro-channel chip, it is characterized in that: comprise two wall bodies and an annular gasket, two wall body clamping annular gaskets form airtight microcavity, wherein a wall body is provided with biomaterial entrance and biomaterial outlet, biomaterial entrance and biomaterial outlet all communicate with airtight microcavity, are equipped with several grooved passages on the outside surface of two wall bodies.
2. biomaterial low temperature according to claim 1 is preserved micro-channel chip, it is characterized in that: the arrangement of set grooved channel parallel and length and micro-channel chip wall body are equal in length.
3. biomaterial low temperature according to claim 1 and 2 is preserved micro-channel chip, it is characterized in that: the xsect of described grooved passage is rectangular, semicircle or triangle.
4. biomaterial low temperature according to claim 1 is preserved micro-channel chip, it is characterized in that: described biomaterial entrance and biomaterial outlet lay respectively on two diagonal angles of biomaterial entrance and biomaterial outlet place wall body.
5. biomaterial low temperature according to claim 1 is preserved micro-channel chip, it is characterized in that: described wall body is silicon chip or sheet metal.
6. biomaterial low temperature according to claim 1 is preserved micro-channel chip, it is characterized in that: the material of described annular gasket is flexible high molecular material.
7. biomaterial low temperature according to claim 6 is preserved micro-channel chip, it is characterized in that: the material of described annular gasket is dimethyl silicone polymer.
8. a biological material low temperature storage device, it is characterized in that: comprise that the biomaterial low temperature described in claim 1～7 any one claim preserves micro-channel chip, also comprise airtight lagging casing, described micro-channel chip is fixed on the central authorities of lagging casing, the lagging casing one side parallel with micro-channel chip is provided with hydraulic fluid entrance, hydraulic fluid outlet, biomaterial entrance and biomaterial outlet, wherein, the position of the position of biomaterial entrance and biomaterial outlet and set biomaterial entrance on micro-channel chip and biomaterial outlet is corresponding and by pipeline, exported and be communicated with the biomaterial entrance arranging on micro-channel chip and biomaterial respectively, hydraulic fluid entrance is corresponding with the position at place, set grooved passage two ends on micro-channel chip respectively with the position of hydraulic fluid outlet.
9. biological material low temperature storage device according to claim 8, it is characterized in that: the middle part of described lagging casing is provided with a dividing plate, the center of dividing plate has the hole that a size is mated with micro-channel chip size, this hole is used for installing micro-channel chip, exports corresponding position be respectively equipped with engraved structure on dividing plate with hydraulic fluid entrance and hydraulic fluid.
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Cited By (3)
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| CN104748605A (en) * | 2015-03-25 | 2015-07-01 | 华南理工大学 | Electric field enhanced nanofluid phase change heat transfer compact type multi-layer microchannel heat exchanger |
| CN111402728A (en) * | 2020-03-24 | 2020-07-10 | 京东方科技集团股份有限公司 | Bent gasket subassembly, flexible O L ED module and O L ED equipment |
| CN117305104A (en) * | 2023-09-04 | 2023-12-29 | 上海交通大学 | Scientific experiment system with multiple physical fields |
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Cited By (4)
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Application publication date: 20140326 |