CN203492656U - Blood bag gradually-freezing preservation device - Google Patents
Blood bag gradually-freezing preservation device Download PDFInfo
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- CN203492656U CN203492656U CN201320456299.2U CN201320456299U CN203492656U CN 203492656 U CN203492656 U CN 203492656U CN 201320456299 U CN201320456299 U CN 201320456299U CN 203492656 U CN203492656 U CN 203492656U
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Abstract
The utility model relates to a blood bag gradually-freezing preservation device. The blood bag gradually-freezing preservation device comprises a main body shell, more than one layer of blood bag containing space and more than two Teflon gradually-freezing bags, wherein the main body shell is provided with an upper cover which can be opened and closed, a blood bag to be frozen and preserved is arranged in the blood bag containing space, the Teflon gradually-freezing bags are contacted with the blood bag to be frozen and preserved, and the Teflon gradually-freezing bag are filled with gradually-freezing agents. According to the blood bag gradually-freezing preservation device, after the gradually-freezing preservation device is assembled, the blood bag to be frozen and the gradually-freezing bags are arranged in a stacked manner and are arranged inside a main body of the blood bag gradually-freezing preservation device; besides, the Teflon gradually-freezing bag is directly contacted with the blood bag to assist the blood bag to achieve the slowly-cooling effect during freezing of the blood bag.
Description
Technical field
The utility model relates to a kind of blood bag cryopreservation device, and particularly a kind of blood bag gradually freezes save set.
Background technology
While preserving cell with freezing mode, must, through the process of one section of slow cooling, avoid fast cooling or unstable cooling to cause ICW in refrigerating process because crystalloid condenses, to produce cell death.Success frozen cell needs two primary conditions, and the one, add cryoprotective agent, the 2nd, with best rate of temperature fall cooling.Conventional cryoprotective agent is dimethyl sulfoxide (DMSO) (DMSO); DMSO is a kind of minimum molecule, can enter in cell by cell membrane, forms size and the speed of ice crystal to slow down ICW; Cell protection is not damaged in stored frozen and course of defrosting, increases cell survival rate.But when DMSO concentration is high, also there is cytotoxicity, therefore need be controlled at 10%(wt when frozen) below, best rate of temperature fall merged to reach the freezing preservation of the best of cell.
General commercially available cell gradually freezes conserving case (Cryo box) for can reach best rate of temperature fall, can in the middle of inner box object-putting layer and shell, fill 100% isopropyl alcohol (isopropyl alcohol) as gradually freezing agent, can decline per minute the temperature of 1 ℃ stablize cooling, minimum reaching-80 ℃, and can maintain for a long time low temperature.Isopropyl alcohol can be assisted and realize slow cooling in the process of frozen cell, utilizes this material can make rate of temperature fall reach and is similar to-1 ℃/min, also can obtain the effect of level and smooth temperature lowering curve in frigorimeter.But isopropyl alcohol has height volatility, and easily absorbs airborne moisture, so after long use, its concentration can decline gradually and affect its temperature control performance.Therefore, the frozen pipe that Ke Luonai (Kelowna) International Technology Co., Ltd produces gradually freezes box, and do not need to add isopropyl alcohol and reach and gradually freeze effect, and due to can fast temperature rising, therefore can shorten the time reusing.But so far, the known commercially available box (the Mr. Frosty producing as Nalgene) that gradually freezes only supplies frozen pipe frozen, there is no the open use of can the frozen blood bag of blood supply bag gradually freezing the Related products such as box.
U.S. cell therapy IPCA (FACT) standard points out, Cord blood must be stored in freeze proof blood bag and preserve, because the seal of freeze proof blood bag is high, can reduce opportunities for contamination.The store method that at present general industry adopts is that the Cord blood that collection is obtained packs in the freezer bag (cryo preserved bag) that for example Pall company (U.S.) produces.Pall freezer bag is a kind of new-type blood bag, comprises two parts with separation, and wherein a part of capacity is 20 milliliters, and another part is 5 milliliters.The latter can be before stem cell transplantation for oneself's amplification, then with the former homologous transplantation to patient, or be used for the treatment of impaired histoorgan.In order to prevent cross pollution, after can separately adding one deck sack, freezer bag outside packs an aluminium alloy box into, then this can being put into controlled rate cooling instrument (computerised temperature-reducing system), to carry out program freezing, and rate of temperature fall is-1 ℃/min.When temperature drops to-80 ℃, Cord blood will be proceeded in liquid nitrogen case, (maintenance≤-190 ℃ of low temperature) long-term preserve (over 1 year) in gaseous state.But controlled rate cooling instrument somewhat expensive, except stem cell being stored and transplanting cost increases, standard value when 1 ℃ of decline per minute is frozen pipe Small Amount freeze-stored cell, though for can be used as a reference value in blood bag during mass storage cell, but best rate of temperature fall has inching because of cell quantity, therefore cell in blood bag post-thaw survival rate lower than frozen pipe in post-thaw survival rate, at present countries in the world rules and regulations blood bag thaw after cell survival rate as reached more than 70%, be one of clearance standard of clinical use.
Utility model content
In view of this, the purpose of this utility model is to provide a kind of blood bag gradually to freeze save set.
For achieving the above object, the utility model provides a kind of blood bag gradually to freeze save set, comprising:
One main body cover, it is provided with one can be for opening, airtight upper cover, and gradually freeze after save set has been assembled and present sealing in described blood bag;
Blood bag more than one deck is inserted space, and described blood bag is inserted space and is equipped with the frozen blood bag of wish; And
Two above Teflons gradually freeze bag, and described Teflon gradually freezes a bag blood bag frozen with wish and contacts, and described Teflon gradually freeze in bag, be filled with can make rate of temperature fall reach-1 ℃/min gradually freeze agent.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, wherein said gradually to freeze agent be isopropyl alcohol.Gradually freeze bag size and take that to coordinate the length and width of this main body inner edge be principle, that wherein filled gradually freezes agent weight (g)/gradually freeze a bag area (cm
2) ratio is preferably 0.7 ~ 0.9, this considers the used better bearing capacity that gradually freezes bag.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, space inserted by wherein said blood bag and described Teflon gradually freezes bag to pile alternately stack mode, is arranged in from the bottom to top the body interior that described blood bag gradually freezes save set.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, the upper and lower layer that wherein said blood bag is inserted space all gradually freezes bag with described Teflon and is connected, and the above and below of the blood bag of inserting can touch respectively described Teflon and gradually freezes bag.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, the main body that the upper cover of wherein said main body cover gradually freezes save set with mode and the described blood bag of pressing is closely sealed, to avoid content pollute and conveniently transport and storehouse.
The preferred version that gradually freezes save set as above-mentioned a kind of blood bag, the inner face periphery of wherein said upper cover is provided with the annular ditch groove closely sealed with the opening periphery of described main body cover.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, the upper cover of wherein said main body cover is closely sealed with the main body that described blood bag gradually freezes save set in the mode fastening, to avoid content pollution and conveniently to transport and storehouse.
The preferred version that gradually freezes save set as above-mentioned a kind of blood bag, wherein said main body cover and upper cover are respectively equipped with the exterior buckle type device of more than one group mutual fastening.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, wherein said main body cover and upper cover are made by resistance to plastics, stationery or the metal of freezing.
The preferred version that gradually freezes save set as above-mentioned a kind of blood bag, wherein said main body cover and upper cover are made by PP plastics (polypropylene).
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, it is the can that can dismantle separately that wherein said blood bag is inserted space, or the space being formed by the dividing plate that can be set up in the bump of this main body cover inwall, described blood bag is inserted space must possess the upper and lower outer surface that can make the blood bag of inserting, and can gradually freeze the height that bag contacts with this Teflon.
As above-mentioned a kind of blood bag, gradually freeze the preferred version of save set, the wherein said bag size of gradually freezing take that to coordinate the length and width of this main body cover inner edge be principle, and that wherein filled gradually freezes agent weight (g)/gradually freeze a bag area (cm
2) ratio is 0.7 ~ 0.9, this considers the used better bearing capacity that gradually freezes bag.
The utility model is alleged " blood bag ", be extensively defined as the freezer bag that wherein includes cell culture.For herein, described cell culture can be in vitro through a large amount of cultured cells cultures, including (but not limited to) stem cell culture, or gathers from the Cord blood of live body.
The blood bag that the utility model provides gradually freezes save set, when this gradually freezes device and has assembled, want to carry out freezing blood bag and this and gradually freeze bag with overlapped way, arrangement is placed in the body interior that this blood bag gradually freezes save set, and this Teflon gradually freezes bag and directly contacts with blood bag, assist blood bag in refrigerating process, to reach the effect of desirable slow cooling; It has and gradually freezes function, and with low cost; And cell culture is placed in this blood bag, gradually freeze save set, can promote a large amount of cells and be stored in the cell survival rate after blood bag thaws.
Accompanying drawing explanation
Fig. 1 is for gradually to freeze the perspective view of save set embodiment main body according to a blood bag of the present utility model;
Fig. 2 is the cutaway view that the of the present utility model one blood bag that completes assembling with Sanming City therapy gradually freezes save set embodiment;
Fig. 3 is the desirable temperature lowering curve (Data Source: Transfus Med Rev. 1997 Jul of the standard of cell freezing process; 11 (3): 224-33);
Fig. 4 A is for gradually freezing save set in freezing preservation method process with the blood bag of arranging assembling in sandwich mode of the present utility model, in the temperature lowering curve that comprises that 2 layers of different sample point of not yet inserting the empty iron clamp of blood bag measure;
Fig. 4 B is for gradually freezing save set in freezing preservation method process with the blood bag of arranging assembling in sandwich mode of the present utility model, in the temperature lowering curve that comprises that 2 layers of different sample point that gradually freezes box (CryoBag) that has been placed into blood bag measure;
Fig. 5 is that the blood bag that alternate manner is arranged assembling gradually freezes save set in freezing preservation method process, in the temperature lowering curve that comprises that 2 layers of different sample point that gradually freezes box (CryoBag) of having inserted blood bag measure;
Fig. 6 is the frozen mesenchymal stem cell (mesenchymal stem cells, MSCs) of freezing method for storing gradually of the blood bag through in the utility model, the cell survival rate figure recording after thawing.
[primary clustering symbol description]
1-blood bag gradually freezes save set; 10-main body cover; 12-upper cover;
20-blood bag is inserted space;
30-gradually freezes bag;
The freezing storage bag of 40-.
Embodiment
Other characteristic of the present utility model and advantage will further be given an example and explanation in the following example, and this embodiment is only as aid illustration, not for limiting scope of the present utility model.
First with reference to Fig. 1 and Fig. 2, describe in detail and gradually freeze the structure of save set 1 and the embodiment of assembling mode as a blood bag of the present utility model.As shown in Figure 1, blood bag of the present utility model gradually freezes save set 1, include that a main body cover 10, blood bag more than one deck are inserted space 20 and two above Teflons (Teflon) gradually freeze bag 30, this main body cover is provided with one can be for opening, airtight upper cover 12, this every one deck blood bag is inserted space 20 can be for inserting two above blood bags, this Teflon (Teflon) gradually freezes bag and is filled with in 30 and gradually freezes agent, and when assembling, be installed in respectively the upper and lower part that every one deck blood bag is inserted space 20, and directly contact with blood bag.
For reaching best controlled-rate freezing, the internal capacity that described main body cover 10 forms be take can for holding at the most 2 layers of blood bag and 3, gradually to freeze bag 30 be principle, that is gradually freeze save set 1 with single blood bag of the present utility model, in better carrying, be limited to 4 blood bags, individual layer blood bag is inserted and in space 20, is inserted 2 blood bags at the most, and the size that this gradually freezes bag 30 with the lucky length and width of this main body inner edge that can coordinate for better.Teflon gradually freezes bag 30 inner fillings to be had and gradually freezes agent (known preferably gradually freeze agent be 100% isopropyl alcohol (isopropyl alcohol)), the ratio that dosage and this gradually freeze bag 30 areas of gradually freezing wherein filled is, weight (g)/gradually freeze a bag area (cm
2)=0.7 ~ 0.9.For instance, the sack inner edge length and width of take are 21 cm x 13.5 cm gradually freezes bag 30, wherein better can filling 100% isopropyl alcohol of 200 grams.
The utility model blood bag gradually freezes the assembling mode example of save set 1: this is gradually frozen to bag 30 precooling more than 2.5 hours in-30 ℃ of refrigerators.The single blood bag that collection is obtained is inserted (this combination is designated hereinafter simply as freezing storage bag 40) in iron clamp, is first temporary in 4 ℃ of refrigerators, after prepared by all freezing storage bags 40, carries out the assembling that the utility model blood bag gradually freezes save set 1.Assembling mode preferably carries out interaction cascading arrangement (as shown in Figure 2) with Sanming City therapy, wherein by of the present utility model gradually freeze bag 30 layered stacks on blood bag 40 and below, and make freezing storage bag 40 be arranged in upper and lower two, gradually freeze between bag 30, the upper and lower faces of freezing storage bag 40 like this all can directly touch and gradually freeze bag 30.In gradually freezing 30, bag, can laterally keep flat the freezing storage bag 40 of individual layer, maximum 2 the freezing storage bags 40 of individual layer, and confirm freezing storage bag 40 all average force in gradually freezing on bag 30, completing, gradually freeze after bag 30 and the mutual storehouse of freezing storage bag, can cover the upper cover 12 of this body housing 10.This upper cover 12 with gradually freeze device body 10 can pressing or the mode of fastening closely close entire.After assembling completes, will gradually freeze save set 10 and be placed in-80 ℃ of refrigerators and reach more than 6 hours, be preferably 6-10 hour, make it be cooled to gradually-80 ℃, can move in liquid nitrogen bucket and preserve.
Cell freezing must make intracellular moisture form vitrifying and solidify through the process of slow cooling, and non-rapid condensation becomes irregular crystallization and injures cell.The iptimum speed that is cooled to crystallization formation (Pre-freeze) is 1 ℃ (1 ℃/min) of temperature decline per minute.(extracts is from Transfus Med Rev. 1997 Jul as Fig. 3 for the desirable temperature lowering curve of its standard (STD); 11 (3): 224-33).
Therefore, further test the utility model is with sandwich overlapped way (hereinafter to be referred as sandwich freezing preservation method), and other arrangement mode assemble gradually freeze device, the temperature lowering curve obtaining, the cell freezing effect of gradually freezing save set 1 to understand the utility model blood bag in temperature-fall period.In sandwich freezing preservation method process, the temperature lowering curve that these 2 layers empty iron clamp of not yet inserting blood bag measures is consistent, and its rate of temperature fall approaches the STD (as shown in Figure 4 A) of 1 ℃ of decline per minute.Blood bag with the assembling of the utility model Sanming City therapy gradually freezes save set 1, in whole frozen process, these 2 layers of blood bag heat release times are short, the heat release temperature difference (Δ T) little (1.6 ~ 2.6 ℃), though rate of temperature fall is slightly slower than STD, but it is good that a large amount of cells are placed in to blood bag preservation effect, show that these 2 layers freezing storage bag 40 cooling situation are similar, cooling slowly, heat release have a narrow range of temperature (shown in Fig. 4 B).And with the direct storehouse of two-layer blood bag, in the upper and lower layer of these two layers of freezing storage bags 40, place the freezing preservation method (as Fig. 5) that gradually freezes bag again, by its temperature lowering curve, can be found, heat release time length and the heat release temperature difference (Δ T) of the liquid crystalization of these 2 freezing storage bags 40 are large, represent that in the utility model, the effect of gradually freezing with sandwich freezing preservation method is better than the freezing preservation method with the direct storehouse of blood bag.
The comparison of cell survival rate (%).Cell is utilized to other cryopreservation methods storehouse freezing preservation method as direct in blood bag and sandwich freezing preservation method, respectively at carrying out cell at-80 ℃, gradually freeze, then be placed in liquid nitrogen and preserve, after taking out and thawing, counting cells in cells frozen storing liquid.Result shows, the MSCs (mesenchymal stem cells) thawing after frozen with Sanming City therapy, has preferably cell survival rate (90.89%), compared to the direct storehouse freezing preservation method of blood bag, has significant difference (Fig. 6).And with regard to the observed result of cellular morphology, MSCs is carried out after thawing to subsides dish to cultivate 3 days, find that these cells, after recovering, still have the characteristic that it pastes growth.
Utilize the utility model gradually to freeze the MSCs that method is gradually frozen, after frozen, thaw, then carry out cell surface antigen evaluation, and in vitro differentiation capability analysis.After cell being dyeed with the specific antibodies with fluorescence, whether by flow cytometer, detecting cell has performance MSC specific markers (markers).
Table 1.
Result in table 1 shows, 3 batches of MSC cells that gradually freeze, measured performance CD105 after thawing, CD90, the cell proportion of CD73 is all greater than 95%, and do not show Negative Cocktail markers (CD45, CD34, CD14 or CD11b, CD79 α or CD19, HLA-DR) cell mass is greater than 98% (being Negative markers performance amount <2%), the antigen performance that result all meets the defined mankind's mesenchymal stem cell of international Society for cellular therapy (ISCT) (MSC) (please refer to Cytotherapy (2006) Vol. 8 according to ISCT to the definite condition of MSC, No. 4, 315-317).
After cell subsides dish, guide and be divided into " skeletonization ", " cartilage " and " fat " cell, and with dyeing identification of cell differentiation situation.The staining method of various cells and colour developing result are shown in following table 2.
Table 2.
The result demonstration of table 2, the MSCs that 3 batches (SW01, SW02, SW03) gradually freezes, after induction differentiation, all has the differentiation capability that is divided into Gegenbaur's cell, cartilage cell and adipocyte.Result all meets the differentiation potential (tri-lineage differentiation capacity) of the defined mankind's mesenchymal stem cell of ISCT.
By above-mentioned cell measurement result, summed up, with other blood bag freezing preservation method comparison, the cryopreservation methods that blood bag of the present utility model gradually freezes save set 1 and uses it to carry out, can provide and be beneficial to the cooling conditions that cell freezing is preserved, make a large amount of cells in blood bag, maintain preferably survival rate (90.89%).And, utilize blood bag of the present utility model gradually to freeze save set 1 and gradually freeze rear frozen MSCs, after thawing, still there is characteristic and the differentiation capability of MSC, all meet the clearance standard of ISCT to clinical use MSC.
The above, be only preferred embodiment of the present utility model, not in order to limit the scope of the claims of the present utility model.Therefore such as use equivalent modifications that the utility model specification and accompanying drawing content do or variation etc., all should in like manner belong in scope of patent protection of the present utility model.
Claims (8)
1. blood bag gradually freezes a save set, it is characterized in that, comprising:
One main body cover, it is provided with one can be for opening, airtight upper cover, and gradually freeze after save set has been assembled and present sealing in described blood bag;
Blood bag more than one deck is inserted space, and described blood bag is inserted space and is equipped with the frozen blood bag of wish; And
Two above Teflons gradually freeze bag, and described Teflon gradually freezes a bag blood bag frozen with wish and contacts, and described Teflon gradually freeze in bag, be filled with can make rate of temperature fall reach-1 ℃/min gradually freeze agent.
2. blood bag according to claim 1 gradually freezes save set, it is characterized in that, described gradually to freeze agent be isopropyl alcohol.
3. blood bag according to claim 1 gradually freezes save set, it is characterized in that, space inserted by described blood bag and described Teflon gradually freezes bag to pile alternately stack mode, is arranged in from the bottom to top the body interior that described blood bag gradually freezes save set.
4. blood bag according to claim 3 gradually freezes save set, it is characterized in that, the upper and lower layer that described blood bag is inserted space all gradually freezes bag with described Teflon and is connected, and the above and below of the blood bag of inserting can touch respectively described Teflon and gradually freezes bag.
5. blood bag according to claim 1 gradually freezes save set, it is characterized in that, the main body that the upper cover of described main body cover gradually freezes save set with mode and the described blood bag of pressing is closely sealed.
6. blood bag according to claim 5 gradually freezes save set, it is characterized in that, the inner face periphery of described upper cover is provided with the annular ditch groove closely sealed with the opening periphery of described main body cover.
7. blood bag according to claim 1 gradually freezes save set, it is characterized in that, the main body that the upper cover of described main body cover gradually freezes save set with the mode that fastens and described blood bag is closely sealed.
8. blood bag according to claim 7 gradually freezes save set, it is characterized in that, described main body cover and upper cover are respectively equipped with the exterior buckle type device of more than one group mutual fastening.
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| CN201320456299.2U CN203492656U (en) | 2013-07-29 | 2013-07-29 | Blood bag gradually-freezing preservation device |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104336005A (en) * | 2013-07-29 | 2015-02-11 | 仲恩生医科技股份有限公司 | Blood bag gradually-freezing preservation device and application method thereof |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104336005A (en) * | 2013-07-29 | 2015-02-11 | 仲恩生医科技股份有限公司 | Blood bag gradually-freezing preservation device and application method thereof |
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