Abstract:
A device for mixing biological specimens contained in flexible storage bags ( 104 ) at controlled temperature, comprising a support ( 105 ) for supporting a storage bag containing a biological specimen to be mixed; means for imparting a displacement to a specimen in a storage bag on the support to mix the specimen; and temperature control means for maintaining the specimens at a controlled temperature during mixing. The means for imparting displacement to a specimen comprises at least one inflatable/deflatable bag ( 102, 103 ) (air bag) that when inflated contacts the surface of a part of a storage bag to progressively squeeze the storage bag and displace the contained specimen into another part of the storage bag.

Description:
FIELD OF THE INVENTION 
       [0001]    The present invention relates to an automated mixing system, and in particular a system capable of mixing safely, smoothly, homogeneously and efficiently biological specimens like whole blood, placenta/umbilical cord blood, bone marrow, apheresis product, or stromal vascular fraction (SVF) contained in a flexible collection, freezing, storage or transfer bag, especially during addition of an organosulfur compound like dimethyl sulfoxide (DMSO) or other biological additives. During the whole process duration, the automated system maintains biological specimens at a stabilized temperature. 
         [0002]    Such procedures are likely to be performed in hospital environments and medical or biological laboratories. Typical types of blood or biological specimen manipulations are: preparation for long-term storage, thawing after long term storage, preparation prior to transplantation, cells culture expansion, adipose tissue manipulations or other similar applications. 
       BACKGROUND OF THE INVENTION 
       [0003]    The role of stem cells in transplantation or regenerative medicine cell therapies is rapidly expanding. Within the field of transplantation, the current therapeutic strategy demands that progenitor cells are cryopreserved for virtually all autologous and many allogeneic transplants. The cryopreservation process is of importance for all types of stem cell collection and over the years, freezing and thawing techniques have proven to be efficient and to have the ability of long-term storage with a high survival rate of biological cells. 
       Cryopreservation Process 
       [0004]    The cryopreservation process consists of storing a biological fluid in a liquid or vapour phase nitrogen at generally −196° C. in mechanical freezers. The freezing rate is controlled and concentrated stem cells are frozen down at a typical rate of 1-2° C./min. 
         [0005]    In order to freeze down and store concentrated cells at the mentioned temperatures, it is necessary to add an organosulfur compound like dimethyl sulfoxide (DMSO) to the biological specimen. In cryobiology, DMSO cryoprotectant is used for preserving organs, tissues, and cell suspensions by preventing freezing damage to living cells. It inhibits the formation of intra and extracellular crystals and hence cell death. 
         [0006]    Without cryoprotectant additives, up to 90% frozen cells would become inactive. In general, 25% DMSO mixture is added to hematopoietic stem cells before cryopreservation. 
         [0007]    In a standardized environment like cord blood banking, or in an hospital environment after an autologous apheresis collection, there is a need to add cryoprotectant before long-term storage in liquid or vapour phase nitrogen. In general, a sterile chilled DMSO is added to the blood bag over the course of 10-15 minutes. While the biological specimen should be maintained at a stable temperature, the DMSO must be added gradually for protecting cells from damaging osmotic stress due to high DMSO concentrations and to prevent an exothermic reaction which occurs as the concentrated DMSO solution mixed with the cell solution. Furthermore, it is important to ensure that DMSO or biological additive is homogeneously mixed with the blood bag or biological specimen during addition to ensure an efficient protective effect during cryopreservation. 
         [0008]    Outside needs for cryopreservation preparation, many applications require additives to be mixed with biological specimens. For example, in cell therapy or research laboratories, often cell concentrated products need to be mixed with culture medium solutions for proliferating cells, or washing or diluting enriched cell products with washing buffers like sodium chloride (NaCl) or Ringer&#39;s lactate solutions. In this setup, again there is a need for ensuring a proper gradual, smooth and homogeneous mixing under controlled stabilized temperature. 
         [0009]    Another possible application is thawing cryopreserved specimens. Today, most allogeneic cord blood units and autologous apheresis units are thawed using a standard water bath technique, which consists of laying under water at 37° C., blood bags pulled out from liquid or vapour phase nitrogen. In this method, water is the thermal transfer vector to the bag, and therefore, has a potential risk of microbiological contamination. It is thus desirable to have a repeatable system able to thaw specimens using a dry-tempering system by maintaining at stabilized 37° C. or predefined temperature, to ensure an efficient thawing, whereby a blood bag is mixed smoothly during the whole process duration. 
         [0010]    Thus, risk of microbiologic contamination is reduced and the product is not subject to operator variability during mechanical agitation of bags inside water. 
         [0011]    Within the field of stromal vascular fraction banking, many steps are necessary for extracting adipose tissue from a patient, digesting and processing the fat until the stromal vascular fraction is concentrated with mesenchymal stem cells and is ready for banking. Within these steps, adipose tissue should in general be washed with washing buffer solution like Ringer&#39;s lactate or sodium chloride, and adipose must be digested at stabilized 37° C. temperature with collagenase enzymes. It would thus be desirable to provide an automated system able to program a stabilized temperature, to pump additives into adipose tissue and to ensure a homogeneous and smooth mixing of biological specimens. 
         [0012]    Nowadays, the mixing process is still very operator dependant and there remains a widespread need for providing an automated system able to ensure a safe, efficient and homogeneous mixing especially when adding biological additives to blood products or other types of biological specimens. 
       State of the Art 
       [0013]    The most common mixing technologies for biological specimens nowadays are shaking techniques or a mechanical arm movement interacting with the biological specimen. 
         [0014]    The Coolmix™ device from Biosafe S. A. is an automated mixing device that allows the preparation of stem cells for cryopreservation. The system uses a mixing mechanism by the intermediary of a mechanical arm moving up and down, thus a vortex movement is created when a surface of the bag is squeezed by the metallic arm. 
         [0015]    The known systems suffer from the limitations that they may produce rapid movements or high gravitational acceleration of cells or manipulated products and there is a risk of mechanical friction or rough shaking of the cells or manipulated products and customization of the devices is difficult. 
       Disclosure of the Invention 
       [0016]    As set out in the claims, the invention proposes an improved automated system for ensuring a safe, smooth, homogeneous and efficient mixing of collection, freezing, storage or transfer bags containing either blood or biological solution, especially during injecting an organosulfur compound like dimethyl sulfoxide (DMSO) or other biological additives. 
         [0017]    According to a main aspect of the invention, there is provided a device for mixing biological specimens contained in flexible collection, freezing, storage or transfer bags (hereinafter “storage bags”) at controlled temperature, comprising:
       a) A support for supporting a storage bag containing a biological specimen to be mixed;   b) Means for imparting a displacement to a specimen in a storage bag on the support to mix the specimen; and   c) Temperature control means for maintaining the specimens at a controlled temperature during mixing.       
 
         [0021]    The device according to the invention is characterized in that the means for imparting displacement to a specimen comprises at least one inflatable/deflatable bag (hereinafter “air bag”) that when inflated contacts the surface of a part of a storage bag to progressively squeeze the storage bag and displace the contained specimens into another part of the storage bag. 
         [0022]    Further aspects of the invention are set out in the claims. 
         [0023]    The invention concerns also the overall mixing apparatus and instrumentation capable of mixing smoothly and homogeneously one small or large bag, or two bags or several bags at the same time. 
         [0024]    The invention provides a mixing system composed of one or several pneumatic air-bags in contact with blood biological specimen bags. Air-based bags are inflated and deflated at several frequencies and with customized profiles depending on the types of blood bags, volume and sensitivity of both products to be mixed. This action results in squeezing the biological specimen over a defined surface with a distributed force. 
         [0025]    The present invention provides hospital environments, medical and biological laboratories an automated system able to mix smoothly specimens, and in contrast with existing shakers guarantee no rapid movement and no high gravitational accelerations of cells or products manipulated. Technique used with present invention is the application of a smooth pressure on a predefined surface which is rather large, which results in creating an efficient vortex effect ensuring a homogeneous mixing of the specimen. 
         [0026]    Squeezing performed by an inflated air-bag ensures a distributed force along the surface area, which results in smooth and homogeneous mixing of the biological specimen, with less risk of mechanical friction or rough shaking of cells or manipulated products. 
         [0027]    Also, the inventive device can accommodate a variety of bag sizes used for manipulations. The present invention covers a wide range of blood or biological bags of different sizes, either by designing pneumatic-air bags accordingly to specific bag sizes needed to be manipulated, or by integrating a set of air-bags in order to cover a wide range of bag sizes. Pneumatic air bags can also be inflated and deflated with a customized profile adapted for specific blood bag sizes. 
         [0028]    Manufacturing techniques of air-based bags are also inexpensive techniques compared to complex shaking or mechanic movement mechanisms. For the air bags, elastomers like thermoplastic polyurethane or polyvinyl chloride (PVC) are generally used and manufactured by conventional thermoplastic methods. 
       One Bag 
       [0029]    Taking as example a freezing bag containing 20 ml of biological specimens, see WO 2009/138966 (Biosafe), the mixing movement is done by pressurizing and depressurizing an air bag having a sturdy silicon membrane with compressed air. 
         [0030]    The function of the mixing bag is to provide a vertical push and pull movement transmitted to the bag to be mixed. By squeezing a distributed force over an extended surface, a narrow path is created which results in creating a good vortex effect ensuring an efficient and homogeneous mixing. Dual compartment bags, by design, have already a narrow path linking smaller and larger compartment. This narrow path also creates the desired vortex effect. 
         [0031]    Air is supplied from a pneumatic system driven by electro-valves and a pneumatic pump. By doing so, it is possible to control the inflating and deflating rate and speed of movement of the air bag, which ensure an optimized mixing of the biological specimens. 
         [0032]    The body of the device typically has an aluminum dish, on which cryopreservation bags or other types of bags are laid. Biological Bags are mechanically constrained by a cover plate containing an air mixing bag. 
         [0033]    An aluminum dish forming a support of the device is fixed on top of a set of Peltier elements dimensioned to provide enough cooling or heating capacity to the device. 
         [0034]    Thermal insulation around the Peltier element is also provided. Heat is then exchanged with the rest of the device via a heat exchanger and an air flow cooling system. 
       Multiple Bags 
       [0035]    The system can be designed to be able to mix two or several cryoresistant bags at the same time. Several pneumatic air-based bags could be inflated or deflated synchronously or asynchronously according to market needs. In order to control the pneumatic system, a central pneumatic pump with several electro-valves is used to control separately each air-bag. 
       Large Bags 
       [0036]    The system is also able to mix a large collection of bags: freezing, storage or transfer bags. The mixing movement is ensured by two or multiple pneumatic air bags. If two vertical air bags are used for pushing and pulling, they could be placed on each side of the biological specimen. By smoothly squeezing one side surface at a time, and by alternating both sides, the fluid will move from one side to the other side of the bag. Thus, the biological specimen mixed with additive is homogeneously mixed during the whole process duration. 
         [0037]    Optional squeezing lips can be used for ensuring creation of a good vortex for large volume bags. Lips might be used to hold tight large bags on a defined line over the bag, virtually creating two compartments linked with a narrow small path, or to narrow the bag width in the middle for example creating a narrow path all the way long. 
     
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
         [0038]    Features, aspects and advantages of the present invention will become more apparent from the following detailed description when taken in conjunction with the accompanying drawings, in which: 
           [0039]      FIG. 1  is a diagram showing the principle of mixing of biological bags by inflating or deflating air bags; 
           [0040]      FIG. 2A  is a schematic vertical cross sectional view showing the principle of mixing a biological specimen, showing the pneumatic bag deflated; 
           [0041]      FIG. 2B  is a schematic vertical cross sectional view showing the principle of mixing a biological specimen showing the pneumatic bag inflated; 
           [0042]      FIG. 3A  is a schematic view showing the homogeneous mixing principle and vortex effect of a small single compartment bag; 
           [0043]      FIG. 3B  is a schematic view showing the homogeneous mixing principle and vortex effect of a small dual compartment bag; 
           [0044]      FIG. 4A  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume blood bag in a stand-by mode with air bags deflated; 
           [0045]      FIG. 4B  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume bag when the left side of the blood bag is pressed by an inflated air bag, creating a vortex and movement effect to the right side; 
           [0046]      FIG. 4C  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume bag when the right side of the blood bag is pressed by an inflated air bag, creating a vortex and movement effect to the left side; 
           [0047]      FIG. 5A  is a schematic view showing the homogeneous mixing principle and vortex effect of a large transfer bag and  FIG. 5B  shows a variant; 
           [0048]      FIG. 6  is a block diagram showing a configuration of the fluid mixing system according to an embodiment of the present invention; 
           [0049]      FIG. 7  is a flowchart illustrating a sequence of the pumping and mixing process of the system according to the present embodiment; 
           [0050]      FIGS. 8A and 8B  show graphs of different inflation/deflation frequencies; 
           [0051]      FIG. 9  is an overall perspective view of an embodiment of a device according to the invention with its support visible as the cover is in an open position; 
           [0052]      FIGS. 10A and 10B  show possible configuration for fitting large and small cryobags on a baseplate; and 
           [0053]      FIG. 11  is a schematic view of a chassis construction. 
       
    
    
     DETAILED DESCRIPTION 
       [0054]    Hereinafter, an embodiment of the present invention will be described in detail with reference to the drawings. In the embodiment described below, an automated mixing system for biological fluids will be explained by way of examples. 
         [0055]      FIG. 1  is a diagram showing the principle of mixing biological bags by inflating or deflating bags with air. As shown in  FIG. 1 , the automated mixing system is composed of a base plate  105  with for example a blood storage bag  104  laid on it. Above, there is a lid  101  supporting a pneumatic bag  102 . When the pneumatic bag  102  is inflated as indicated by  103 , the pneumatic bag applies homogeneous pressure on the blood bag  104 , when the pneumatic bag  102  is deflated it is not in contact with the blood bag  104 . 
         [0056]      FIG. 2A  is a schematic vertical cross sectional view showing the principle of an automated mixing system using a pneumatic bag. Here, a base plate  212  has a biological specimen (blood)  216 , 217  laid on it. When cover  211  is closed, a squeezing lip  215  creates a narrow path  218  for the biological fluid. The blood bag can be decomposed in three parts, a small reservoir  216 , the narrow path  218  and a large reservoir  217 . 
         [0057]    In the illustrated position, pneumatic air bag  214  is deflated and not in contact with the blood bag. This happens when the system is at stand-by or after an inflation phase. 
         [0058]    Pneumatic bag  214  is controlled by ambient or compressed air coming from a pneumatic system connected via a pneumatic tube  213 . 
         [0059]      FIG. 2B  is a schematic vertical cross sectional view showing the principle of mixing a biological specimen by inflating the pneumatic bag  214 . 
         [0060]    To reach this position, compressed air supplied from a pneumatic system is driven through the air tube  221  with the aim of inflating the air bag  222 . Once the air bag is inflated, the membrane of the air bag is in contact with the small reservoir of the blood bag  225  and distributed pressure is applied homogeneously on the small reservoir surface  223 . 
         [0061]    Biological fluid moves to the large volume reservoir  224  with an increased pressure due to the narrow path  226 . A vortex effect will ensure a good and homogeneous mixing due to the narrow path, to the bag shape and the squeezing lip design. 
         [0062]      FIG. 3A  is a schematic view showing the homogeneous mixing principle and vortex effect of a small single compartment bag  302  laid on a base plate  301 . The inflating/deflating air bag  305  is virtually seen from above as indicated in dotted lines. 
         [0063]    When a small single compartment bag needs to be mixed, a squeezing lip  306  is necessary and creates a narrow path  304 , virtually creating a small and large reservoir on both sides. When the air bag  305  is inflated and in contact with the virtual small reservoir, fluid moves to the large reservoir though the narrow path  304  and a vortex effect  303  is created. This vortex effect ensures an efficient biological mixing. 
         [0064]    This vortex effect is important to ensure a homogeneous and smooth mixing when the specimen is mixed, and especially while an additive is pumped into the bag  302  by an entry tube  307 . 
         [0065]      FIG. 3B  is a schematic view showing the homogeneous mixing principle and vortex effect of a small dual compartment bag  312  laid on a base plate  311 . An inflating/deflating air bag  315  is virtually seen from above as indicated in dotted lines. 
         [0066]    When a small double compartment bag needs to be mixed, by construction of the bag a narrow path  314  already exists and the squeezing lip  316  is not necessary. When the air bag  315  is inflated and in contact with the small reservoir, fluid moves to the large reservoir through the narrow path  314  and a vortex effect  313  is created. This vortex effect is important to ensure a homogeneous and smooth mixing when the specimen is mixed, and especially while an additive is pumped into the bag  312  by an entry tube  317 . 
         [0067]      FIG. 4A  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume blood bag. The universal base plate is represented by  401 , and the cover by  400 . 
         [0068]    The large volume blood bag is virtually separated into a left  406  and right  407  reservoir, by the intermediary of a narrow channel  409  created by squeezing lips  408  on the base plate  401  and cover  400 . The two virtual volume reservoirs may have equivalent or different volumes depending on the bag shape or the volume to mix. 
         [0069]    A pneumatic system is composed of two left  404  and right  405  pneumatic bags and by two left  402  and respectively right  403  air tubes connected to a pneumatic device. In the illustrated position, the air bags  404 ,  405  are not in contact with the blood bag. 
         [0070]      FIG. 4B  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume bag when the left side of the blood bag  414  is pressed by an inflated air bag  412 , creating a movement of the fluid and a vortex effect from the left reservoir  414  toward the right reservoir  413 . In this configuration, the compressed air comes from the air tube  410  and inflates the left air bag  412 . The right air bag  415  is deflated and is not in contact with the blood bag. 
         [0071]      FIG. 4C  is a schematic vertical cross sectional view showing the homogeneous mixing principle of a large volume bag when the right side of the blood bag  424  is pressed by an inflated air bag  423 , creating a movement of the fluid and a vortex effect of the fluid from the right reservoir  424  toward the left reservoir  425 . In this configuration, the compressed air comes from the air tube  421  and inflates the right air bag  423 . Left air bag  422  is deflated and not in contact with the blood bag. 
         [0072]      FIG. 5A  is a schematic view showing the homogeneous mixing principle and vortex effect of a large transfer bag  502  laid on base plate  501 . One inflating/deflating air bag surface  503  is virtually seen from above, as indicated in dotted lines. In order to create two virtual separate reservoirs, squeezing lips  506  are necessary and create a narrow path  504  for the fluid. When one air-bag is inflated with air, it pushes the equivalent volume of fluid on the other reservoir via the narrow path  504 . During the inflating phase of one side air-bag  503 , fluid passing through the narrow path  504  creates a vortex effect  505  on the other side of the blood bag. Once the air bag  503  is completely inflated on one side, it deflates and the other side air bag inflates creating the same vortex effect on the other side of the blood bag. An alternate movement ensures a homogeneous mixing of the blood bag. This vortex effect is important to ensure a homogeneous mixing especially while an additive is pumped into the bag by a connective tube  507 . 
         [0073]      FIG. 5B  is a schematic view showing a variant of the creation of a vortex effect on a large transfer bag. This vortex effect is created by narrowing the path between two bag sides all the width of the bag. The base plate  511  has the large blood bag  512  laid on it. One inflating/deflating air bag surface  513  is virtually seen from above as indicated in dotted lines.. 
         [0074]    In order to create two virtual separate reservoirs, squeezing lips  514  form a slightly tight path between two bag sides and narrow fluid path  516 . When one side air-bag is inflated, it will be in contact with a side surface, pushing the fluid on the other reservoir via the narrow path  516 . During the inflating phase of one surface  513 , fluid in movement via the narrow path will create a vortex effect on the other side of the blood bag  515 . As in  FIG. 5A , a vortex effect is created when one side is inflated, and an alternative movement ensures a homogeneous mixing of the blood bag  512 . This vortex effect is important to ensure a homogeneous and smooth mixing when the specimen is mixed, and especially while an additive is pumped into the bag by a connective tube  517 . 
         [0075]      FIG. 6  is a block diagram showing a configuration of the fluid mixing system according to the present embodiment. As show in  FIG. 6 , the system is composed of a set of pneumatic air bags of different sizes  110 , and an automated system  100  controlling air flow in the pneumatic bags with appropriate electronics. 
         [0076]    A versatile platform with a set of several air bags is used in order to cover wide shapes and volume range of blood bags. Air bags A1  111  and A2  112  have the same size and are used to mix two small compartment bags simultaneously. Air bags B  113  and C  114  are two larger pneumatic bags able to mix both sides of a large blood bag. 
         [0077]    The main control system  100  ( FIG. 6 ) includes a power supply  120  for supplying electric power to the main CPU and to the system, a Central Processing Unit (CPU)  121  and memory  126  for controlling and monitoring the mixing system. This includes a pneumatic system with solenoid valves  115 - 116 - 117 - 118  and a pneumatic pump  124 , electronic drivers  122  and feedback pressure sensors  125 , thermal control  134  with driver  133  and feedback thermal sensors  135 , and finally peristaltic pumps  131 - 132  and driver  130  for adding additives in blood bags. 
         [0078]    The pneumatic system is controlled by the CPU  121  which determines which bag to inflate or deflate and with which frequency and profile. Instructions provided to the CPU  121  are stored in memory portion  126 . A first action is the control of pump  124  via a driving circuit  122  for generating compressed air for the air-bags. Several valves  115 - 118  are controlled via a driving circuit  123  and have the function to inflate or deflate each bag separately. Valve  118  is the main valve controlling the air flow to all air-bags. Solenoid valve  115  controls simultaneously air bags A1  111  and A2  112 . Those two bags are inflated or deflated at the same time for mixing two small bags synchronously. Solenoid valves  116  and respectively  117  control two larger air-bags  113 ,  114  separately for mixing both sides of a large volume blood bag. An alternate movement ensures the mixing of large bags. A pressure sensor  125  constantly monitors the pneumatic system and provides information to the CPU  121  for control management. 
         [0079]    A thermal control system ensures a stabilized temperature for biological specimen bags. It is composed of a thermal control  134  driven by electronics  133 . Temperature sensor  135  provides information to the CPU  121  for controlling the thermal system. 
         [0080]    Two peristaltic pumps  131 - 132  are also implemented in the system. They are controlled by pump driving circuit  130 , and are used for pumping additive fluids into blood bags. Two peristaltic pumps are necessary when two volume bags are used simultaneously. 
         [0081]      FIG. 7  is a flowchart illustrating a flow of the pumping and mixing process of the system according to the present embodiment. The program according to this flowchart is pre-stored in a memory portion  126 , and the mixing process is carried out as CPU  121  reads out this program from memory portion  126  and executes instructions sequentially. 
         [0082]    As shown in  FIG. 7 , when power is turned on and an application starts, the system is initialised at S 100 . When the system is ready to execute instructions, a first action is to stabilize temperature at a predetermined value at S 101 . Once temperature is stabilized, and a biological bag is correctly inserted in the device, the mixing process S 102  as well as the pumping of the biological additive can start. During the mixing process, air bags is/are constantly inflated at S 103  and deflated at S 104 . Bag inflation is managed by compressed air and stops when a pressure threshold reaches a prescribed level. Bag deflation is managed by an exhaust valve and stops when a second pressure threshold is reached. 
         [0083]    During the mixing and pumping phase, there is a control S 105  checking if a desired volume of additive or a limit of time has been reached. If limitations have not been reached, the process continues and air bag inflates S 103  and deflates S 104  sequentially. 
         [0084]    At the end of the process, sufficient additive fluid has been added to the blood bag or a limit of mixing time has been reached. In this state S 106 , peristaltic pumps  132 ,  132  and bag mixing stop functioning and then the process ends. 
         [0085]    Typical examples of bag mixing are described as follows. 
         [0086]    A small volume bag (e.g. 87×66 mm) containing a biological specimen (e.g. 20 ml) is mixed with a small surface pushed and pulled by an air bag. As the surface in contact is small, the movement needs to be dynamic and repetitive. An average frequency of 0.5 Hz, or one mix every two seconds is adequate. The air bag is inflated via a pneumatic system, and once a maximum pressure of say 300 mBar is reached, a leak valve deflates rapidly the air bag. Then the system repeats the same cycle every two seconds as shown in  FIG. 8A . 
         [0087]    For mixing a typical large volume of about 100 millilitres of biological specimen in a large bag (measuring e.g. approx 240×145 mm), a slower movement is needed with lower pressure. A typical average frequency of 0.1 Hz, or one mix every ten seconds is adequate. The air bag is inflated via a pneumatic system, and once a maximum pressure of 100 mBar is reached, a leak valve deflates rapidly the air bag. Then the system repeats the same cycle every ten seconds, as shown in  FIG. 8B . 
         [0088]      FIG. 9  shows an embodiment of the device according to the invention with its cover  930  open. The device comprises a chassis  920  on which a base plate  901  is mounted. The base plate  901  is adapted for receiving two small or one large cryobags and has a permanent central lips  915  for constricting the middle part of a large cryobag. On the front of the chassis  920  is a touch screen  922  for controlling operation of the device. Also visible are peristaltic pumps  940  for supplying additives during operation. 
         [0089]    The cover  930  is pivotally mounted on the chassis  920  by a hinge  931 . The cover/chassis are provided with means for locking the cover  330  in a closed position on chassis  320  to prevent the cover from opening/raising as a result of inflation of an air bag  952 / 933  during mixing. These locking means can be manually or automatically activated to lock and unlock before and after mixing. 
         [0090]    As shown, in this example, the cover  930  supports two sets of large  932  and small  933  air bags. The two large air bags  932  and small air bags  933  fit on either side of the permanent lip  915  on the support  901 . Between each large air bag  932  and small air bag  933  on the cover  930  is a space  935  for fitting removable lips that can be fitted and removed by the operator and that conveniently can be magnetically held in place against the metal cover  930 . 
         [0091]    Behind the chassis  920  is an upstanding frame with two posts  950  whose height is just greater than the top of cover  930  when open, as shown. 
         [0092]      FIGS. 10A and 10B  show possible configurations for fitting large and small cryobags on a given baseplate  1001  which measures for example 240×145 mm. 
         [0093]    As shown in  FIG. 10A  a single large cryobag  1002  can cover practically the entire baseplate  1001 . In this case, during mixing the cryobag will be divided across part of its middle by the permanent lip  915  ( FIG. 9 ). 
         [0094]    As shown in  FIG. 10B , two small cryobags  1003  and  1004 , measuring for example 87×64 mm, can fit on the baseplate  1001 . The small cryobag  1003  is undivided so in this case the operator will fit a magnetic removable lip at  935  ( FIG. 9 ) to form a constriction across part of the width of the small cryobag  1003 . On the other hand, the small cryobag  1004  already has an integrated division where its two faces are welded together, so there is no need to fit a removable lip. 
         [0095]      FIG. 11  illustrates a side view of the inside of the lower part of the chassis  1120 . The front of the device is to the left of  FIG. 11  and its rear to the right. The chassis  1120  is mounted on legs  1122  and  1124  on unequal length providing the device with an inclination of a few degrees, with the front lower. 
         [0096]    On top of the chassis  1120  is an inclined baseplate  1101  resting on two Peltier elements  1126  which in turn rest on a reticulated/slotted metallic heat sink  1128 . The heat sink  1128  is placed over a fan  1130  that when operated removes warm air from the heat sink. Air enters from below and leaves from the sides. The desired temperature for the baseplate  1101  and hence for specimens being mixed can be set by the operator. 
         [0097]    Using the touch screen  922  the operator can also set the time of mixing and the frequency of inflation/deflation as well as the possible supply of an additive.