PORTABLE AIR BLAST SYSTEM FOR HOMOGENEOUS AND REPRODUCIBLE FREEZING AND THAWING OF BIOLOGICAL MATERIALS

Abstract

This disclosure relates to a system and method to improve the uniformity of the heat transfer coefficient of containers filled with biological materials during the freezing and thawing process, in particular a system that is easily transported and incremented to other conventional freeze-thaw methods and equipment. The system includes a portable air blast system configured to receive a container filled with biological materials and to be placed in a cooling or heating chamber, for homogeneous and reproducible freezing and thawing of biological materials. This disclosure also relates to a method of freezing and thawing biological solution inside a container by using the portable air blast system according to any of the previous claims 1-12, comprising: obtaining a container filled with biological solution; placing the container into the vent enclosure and on the stand of the air blast system; placing the air blast system with the container into a heating or cooling chamber; activating the air blast system by activating the fan in the air blast system.

Claims

1. A portable air blast system for freezing and thawing biological solution inside a container comprising: a vent enclosure with walls; a stand configured to receive a container with side walls; and at least one fan, wherein the fan forces air to pass through the vent enclosure for uniform heat transfer coefficient in the side walls of the container, wherein the distance between the walls of the vent enclosure and the side walls of the container is substantially constant to ensure similar vertical airflow velocity around all side walls of the container.

2. The portable air blast system according to claim 1, further comprising a controller for controlling the vertical airflow velocity.

3. The portable air blast system according to claim 1, further comprising a fan enclosure.

4. The portable air blast system according to claim 1, wherein the distance between the stand and the vent enclosure ranges from 1 cm to 10 cm, and wherein the distance between the walls of the vent enclosure and the side walls of the container ranges from 1 cm to 10 cm and is approximately constant to ensure similar vertical airflow velocity around all side walls of the container.

5. The portable air blast system according to claim 1, wherein the stand further comprises a solid base and pins for connecting the stand to the vent enclosure.

6. The portable air blast system according to claim 1, wherein the stand is configured so that the position of the container is maintained in the center of the vent enclosure.

7. The portable air blast system according to claim 1, wherein the distance between the stand and the vent enclosure ranges from 2 cm to 5 cm.

8. The portable air blast system according to claim 1, wherein the distance between the walls of the vent enclosure and the side walls of the container ranges from 1 cm to 3 cm and is approximately constant to ensure a similar vertical airflow velocity around all side walls of the container.

9. The portable air blast system according to claim 1, wherein the vertical airflow velocity ranges from approximately 0.5 m/s to approximately 20 m/s.

10. The portable air blast system according to claim 1, wherein the system is made of a rigid material.

11. The portable air blast system according to claim 1, wherein the fan is positioned within the fan enclosure.

12. The portable air blast system according to claim 1, wherein the fan is an axial fan.

13. A method of freezing and thawing biological solution inside a container by using the portable air blast system according to claim 1, comprising: obtaining a container filled with biological solution; placing the container into the vent enclosure and on the stand of the air blast system; placing the air blast system with the container into a heating or cooling chamber; and activating the air blast system by activating the fan in the air blast system.

14. The method of freezing and thawing biological solution inside a container according to claim 13, further comprising placing more than one air blast system into a chamber.

15. The method of freezing and thawing biological solution inside a container according to claim 13, wherein the position of the container is maintained in the center of the vent enclosure.

16. The method of freezing and thawing biological solution inside a container according to claim 14, wherein each air blast system is placed at a distance from each other to ensure that the air outside the air blast system vent has a velocity ranging from 0.05 m/s to 1 m/s.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0085] These and other objects, features and advantages of the disclosure will be evident from the following detailed description when read in conjunction with the accompanying drawings.

[0086] For an easier understanding of the disclosure the attached drawings are joined, which represent preferred embodiments of the disclosure that, however, are not meant to limit the object of the present application.

[0087] FIG. 1 shows the time-temperature profile inside a bottle during the freezing process when using blast freezer (solid line) or static freezer (dashed line).

[0088] FIG. 2a is a cross-section view of a bottle 50 frozen in an air blast freezer with a lateral fan. This cross-section view shows the heterogeneity of the ice growth 501.

[0089] FIG. 2b is a lateral view of a bottle 50 frozen in an air blast freezer with a lateral fan. This lateral view shows the lateral deformation 502 of the bottle 50.

[0090] FIG. 3 is a front view of a portable air blast system 10 configured according to the present disclosure.

[0091] FIG. 4 is a schematic cross-section view of a portable air blast system 10 with a container 50 and an ice crust attenuator device 60. The air blast system is configured according to the present disclosure.

[0092] FIG. 5 is a perspective exploded view of a portable air blast system 10 comprising a stand 20, a vent enclosure 30 and a fan enclosure 40. The air blast system is configured according to in the present disclosure.

[0093] FIG. 6 is a partial exploded and elevated view of a portable air blast system 10 configured according to the present disclosure.

[0094] FIG. 7 is an exploded schematic cross-section view of a portable air blast system 10 with a container 50 and an ice crust attenuator device 60. The air blast system is configured according to the present disclosure.

[0095] FIG. 8 is an exploded schematic cross-section view of a portable air blast system 10 with a deformable container 70 inside a rigid shell 701. The air blast system is configured according to the present disclosure.

[0096] FIG. 9 shows the time-temperature profile inside a bottle during the freezing process in a static freezer using (solid line) or not using (dashed line) a portable air blast system 10.

[0097] FIG. 10a is a cross-section view of a bottle 50 frozen in static freezer using a portable air blast system 10. The air blast system is configured according to the present disclosure. The cross-section view shows a planar ice surface 504.

[0098] FIG. 10b is a cross-section view of a bottle 50 frozen in static freezer, showing the ice crust formation and the “pyramidal” shape 501.

[0099] FIG. 11 is a perspective view of multiple portable air blast systems 10 placed in a cooling or heating chamber 80. The air blast system is configured according to the present disclosure.

[0100] FIG. 12 is an exploded section view of a portable air blast system 10 designed in a modular assembly, configured according to the present disclosure.

DETAILED DESCRIPTION

[0101] As described above, the process of freezing and-thawing of biological materials poses several challenges; the lack of homogeneity associated with the freezing and thawing phenomena is one of the main problems. Many variables contribute to freezing inconsistency, the major issue relates to the different freezing rates experienced by multiple containers within a single batch of product. Currently, the freezing process involves placing the bottles and/or carboys comprising the biological materials in a conventional upright or chest freezers, or blast freezers and then allowing the product to freeze. The freezing rate and product quality is dependent on the freezer capacity, freezer load configuration, space between containers, container size, container shape, and airflow properties inside the freezer. When using blast freezers, although the freezing time is faster than using a static freezer (FIG. 1), freezing heterogeneity occurs and this is mainly due to the properties of the airflow (direction and velocity). Depending on the load and configuration of the freezer, this heterogeneity might be higher. In addition to the heterogeneity in the freezing rates between containers, a very important aspect is the heterogeneity of the ice growth 501 in the container 50 (FIG. 2a). This heterogeneity in freezing rates leads to the problem of cryoconcentration and possibly also deformation 502 or rupture of the container 50 (FIG. 2b) due to uncontrolled growth of ice and increased internal pressure. Therefore, it is crucial to have a system and method to freeze and thaw biological materials in a homogeneous and reproducible manner, ensuring controlled ice growth and avoiding the internal pressure build-up.

[0102] Another problem relates to the existence of different freezing and thawing equipment used in several sites of the same biopharmaceutical company. As such, it is desirable to have a portable system, simple for use in different locations, in order to achieve homogeneous and reproducible freezing and thawing of biological materials and can be used in conjunction with existing equipment. Moreover, having a portable system enables the shipping of the air blast system together with any product thus allowing accurate thawing in another location which may have lesser technical capabilities. This ensures the quality of the product and reproducibility of the process throughout the supply chain even when using different equipment.

[0103] The present disclosure describes a system and method that enables improvement in the uniformity of the heat transfer coefficient of the external surface of containers filled with biological materials during the freezing and thawing process, while preventing damage or rupture of the containers. The portable system disclosed allows any chamber to be used for freezing or thawing, and also significantly decreases the time required to freeze or thaw a biological material in a container.

[0104] In an embodiment, the system is configured to receive a container filled with biological materials for freezing and thawing. The system includes a portable air blast system 10 configured to receive a container 50 filled with biological materials. Said portable air blast system 10 is configured to be placed in a cooling or heating chamber, for homogeneous and reproducible freezing and thawing of biological materials, respectively. The main purpose of said portable air blast system is to improve the uniformity of the heat transfer coefficient in the walls of the container, achieving similar freezing/thawing rates even using different cooling or heating chambers. (See FIGS. 3 to 8 for example illustration)

[0105] In an embodiment, the portable air blast system 10 comprises a stand 20, a vent enclosure 30 and a fan enclosure 40. (See FIGS. 3 to 8 for illustrations) The portable air blast system is made of a rigid material, such as plastic, polymer, or other material with high rigidity.

[0106] In an embodiment, the stand 20 is designed to receive a container 50 and to maintain the container's position in the center of the vent enclosure 30, in order to achieve a uniform distribution of air on all the side walls of the container. The stand 20 may have several supports 202 to receive the container 50, designed accordingly to the container to be used with. Said stand 20 may have a solid base 201 and pins 203 to connect the stand 20 to the vent enclosure 30 and to maintain a distance between the stand 20 and the vent enclosure 30. The distance between the stand 20 and the vent enclosure 30 ranges from 1 cm to 10 cm, preferably ranging from 2 cm to 5 cm, this is to ensure that air is distributed into the vent 302. When the portable air blast system 10 is placed inside an air blast chamber with lateral ventilation, the stand 20 may have an opening below, to ensure that air is uniformly distributed in the vent 302. In another embodiment, the container 50 can be maintained in the center of the vent enclosure 30 by a support claw connected to the vent enclosure 30. (See FIG. 5 for illustration)

[0107] In an embodiment, the portable air blast system 10 comprises a vent enclosure 30 designed to create a vent 302, in order to obtain a vertical velocity field that is substantially identical around all side walls 503 of the container. To ensure similar vertical velocity field around all side walls 503 of the container, the distance between the walls of the vent enclosure 301 and the side walls 503 of the container is substantially constant and comprised between 1 cm and 10 cm, preferably between 1 cm and 3 cm. The vent enclosure 30 may decouple the vertical airflow that passes inside the vent 302 of the downward flow that passes outside. The air velocity inside the vent 302 should preferably be higher than the double of the outside air downward velocity. In a preferred embodiment, the air within the vent 302 has a velocity ranging from approximately 0.5 m/s to approximately 20 m/s, preferably ranging from approximately 1 m/s to approximately 10 m/s, and more preferably from approximately 2 m/s to approximately 8 m/s. (See FIG. 7 for illustration)

[0108] In an embodiment, the portable air blast system 10 comprises a fan enclosure 40 with a fan 402 to force air in the chamber or room to pass through the vent 302 in order to improve the uniformity of the heat transfer coefficient on the container's walls during the freezing and thawing process. In a preferred embodiment, the fan 402 is located at the top of the vent enclosure 30, above the container, by using an insulating support 401. In an embodiment, a fan or a blower is used, preferably a fan 402 is used, more preferably an axial fan is used. In another embodiment, batteries could be used to power on the fan, and can be packed inside the insulating support 401. In another embodiment, the velocity of the fan 402 may also be controlled to conveniently increase or decrease the heat transfer for different stages of the process, for sensitive biological materials or for scale-down purposes. In an embodiment, said fan 402 is suitable for use in a cryogenic environment. In another embodiment, more than one fan 402 can be used to force the air to pass through the vent 302, for example 2 or 4 fans juxtaposed horizontally or vertically, or one at the top and another at the bottom of the vent enclosure. (See FIGS. 5 to 7 for illustration)

[0109] In an embodiment, a temperature probe is located at one or more points within the portable air blast system and inside the container. The temperature probe provides an indication of the temperature of the airflow at a particular location inside the portable air blast system and indicates the time-temperature profile of the freezing/thawing of the biological material inside the container. Temperature probe may comprise a thermocouple, a thermistor, or other conventional temperature sensing devices suitable for use in a cryogenic environment.

[0110] In another embodiment, an air velocity probe is located at one or more points of the portable air blast system, to provide information about the airflow velocity inside the portable air blast system at a particular location. Air velocity probe comprises an anemometer, pitot tube, or other conventional sensing devices suitable for use in a cryogenic environment.

[0111] In an embodiment, the container 50 configured to be filled with biological materials can be in several shapes and structural characteristics, such as bottles or carboys. Preferably, said container 50 should maintain its shape when empty and do not significantly deform when filled with product. Said container 50 can be made of a rigid and biocompatible material to promote compatibility with biological materials. The materials can be, for instance, glass, polyethylene terephthalates, polycarbonate, polytetrafluoroethylene, polyethylene, polyesters, polyamides, polypropylenes, ethylene-vinyl alcohol copolymer, polyvinylidenefluoride, polyvinylchlorides, copolymers, and mixtures or laminates that comprise the abovementioned. Said container 50 may vary in size and volumetric capacity. In a preferred embodiment, the container 50 has a volumetric capacity ranging from approximately 1 mL to approximately 20 L, preferably ranging from approximately 100 mL to approximately 10 L. Said container 50 configured to be filled with biological materials may comprise a head-space region, and one cap with at least one port with tubing for aseptic filling and venting purposes.

[0112] In an embodiment, the container 50 may also comprise an ice crust attenuator device 60 configured for attachment to the head-space of the container. The main purpose of the ice crust attenuator 60 device is to prevent the formation of the ice crust that leads to increased pressure inside the containers, and consequently resulting in their damage.

[0113] In another embodiment, the container may also comprise a scale-down device that mimic large-scale containers for development studies and optimization of the freezing-thawing process. In this embodiment using the scale-down device, the air velocity in the vent may be conveniently adjusted to control the average heat transfer coefficient, for example to match that of the large-scale container.

[0114] In another embodiment, deformable containers 70 may also be frozen or thawed with the portable air blast system 10, in this case by placing the deformable container 70 inside a rigid shell 701. The rigid shell is preferably made of a material with low heat resistance such as a metal (for instance, stainless steel, aluminum or copper). Said deformable container 70, such as bags, may deform when filled with product, and can be made of a biocompatible polymeric material to promote compatibility with biological materials. The biocompatible polymeric materials may be, for instance, ethylene-vinyl acetate copolymer, ethylene-vinyl alcohol copolymer, polytetrafluoroethylene, polyethylene, polyesters, polyamides, polypropylenes, polyvinylidenefluoride, polyurethanes, polyvinylchlorides, and copolymers, mixtures or laminates that comprise the above. The deformable container 70, may vary in size and volumetric capacity. In a preferred embodiment, the deformable container 70 has a volumetric capacity ranging from approximately 10 mL to approximately 20 L, preferably ranging from approximately 10 mL to approximately 1 L. (See FIG. 8 for illustration)

[0115] In the present disclosure, biological materials may comprise protein, amino acid and peptide formulations, DNA, RNA and nucleic acid solutions, cell suspensions, tissue suspensions, cell aggregates suspensions, cell growth media, serum, biologicals, blood products, preservation solutions, fermentation broths, and cell culture fluids with and without cells, mixtures of the above and their fragments.

[0116] The portable air blast system 10, depicted in FIGS. 3 to 7, has particular relevance in a common freezing or thawing process when placed directly in a cavity of a chamber. Said chamber may or may not have convection and may be cooled or heated in order to freeze or thaw. Through the use of the fan in the portable air blast system, the air from the cooled or heated chamber will be directed parallel to the walls of the container, without interference from the other adjacent containers, ensuring that the freezing-thawing of each container is not influenced by the others. The system herein disclosed allows any chamber to be used for freezing or thawing, taking advantage of pre-existing cooled/heated air in the chamber. Moreover, the system herein disclosed is designed to be portable, it is lightweight and compact, preferably weighing less than 20 kg or by not increasing the weight and size of the filled container by more than 100%. This shipping between sites can be done easily and can be used in any chamber or room thus ensuring homogeneous, reproducible and faster freezing and thawing of biological materials.

[0117] Further examples are discussed in detail below with regard to the use of the disclosed portable air blast system to freeze an aqueous solution in a container.

[0118] In an embodiment, for example, the portable air blast system 10 was used to freeze a volume of 1.8 L of a 5% (m/V) sucrose aqueous solution in a Polyethylene terephthalate (PET) bottle of 240 (h)×120 (w)×120 (d) mm of dimensions. The test was performed with an ice crust attenuator device 60 as described above. In one experiment, the bottle was placed directly inside an ultra-low freezing chamber with the temperature setpoint at −80° C. and the bottle was allowed to freeze. In another experiment, the bottle was frozen inside the portable air blast system and placed in an ultra-low freezing chamber with the temperature setpoint of −80° C. FIG. 9 illustrates the time-temperature profiles inside the bottle in both experiments, with and without the portable air blast system 10. It can be observed that when using the portable air blast system 10, the freezing process is faster (FIG. 9, solid line). It takes about 200 min to achieve the temperature of −30° C. inside the bottle with the use of the portable air blast system, but it took about 435 min to reach the same temperature without the portable air blast system (FIG. 9, dashed line). Moreover, as shown in FIG. 10, the ice crust formation and the “pyramidal” shape 501 on the top of the liquid are avoid when using the portable air blast system and the ice crust attenuator device, leading to a planar ice surface 504. Therefore, these results show that the system disclosed can improve the freezing process, decreasing the freezing time by improving the heat transfer coefficient on the container's walls during the freezing process while avoiding ice crust formation (FIG. 10a), consequently preventing damage of the containers.

[0119] In another embodiment, a plurality of portable air blast systems 10 could be combined in a single chamber 80. (See FIG. 11 for illustration) In a preferred embodiment, each container has a portable air blast system with one fan above the container. In a preferred embodiment, there is a distance between the several portable air blast systems to ensure that the air outside the vent has a velocity ranging from approximately 0.05 m/s to approximately 1 m/s, preferably ranging from approximately 0.1 m/s to approximately 0.5 m/s.

[0120] In another embodiment, the portable air blast system 10 is be made of modular segments attachable to each other. The portable air blast system 10 comprises the possibility of interchangeable modules for different functions, to receive different containers and devices. The modular assembly of the portable air blast system 10 opens up the possibility of transforming and adapting the system to different scenarios. Different modules can be added or changed to increase the dimensions of the system, to conveniently adapt to different containers, to change the air flow profile and to receive different devices such as probes, batteries, electronics, etc. The modular assembly also allows for ease of transportation, manufacturing, parts replacement and in place assembly. (See FIG. 12 for illustration)

[0121] In another embodiment, the system also includes a trolley to allow quick loading of the chamber with a portable air blast system, or a plurality of portable air blast systems.

[0122] In another embodiment, the container is agitated during thawing to mix the biological solution. This can be achieved by using a platform configured to receive the portable air blast system, that can provide rotation, rocking, shaking, vibrations or other forms of mechanical motion to induce the convection of the liquid inside the container.

[0123] Another aspect of this disclosure relates to a method to improve the uniformity of the heat transfer coefficient of containers filled with biological materials during the freezing and thawing process, using the previously described system, comprising the steps of: (1) providing a cooling or heating chamber; (2) providing at least one portable air blast system; (3) placing the container in the center of the portable air blast system; (4) placing the portable air blast system inside the cooling or heating chamber; and (5) turning on the fan until the biological material completely freezes or thaws.

[0124] The term “comprising” whenever used in this document is intended to indicate the presence of stated features, integers, steps, components, but not to preclude the presence or addition of one or more other features, integers, steps, components or groups thereof.

[0125] It will be appreciated by those of ordinary skill in the art that unless otherwise indicated herein, the particular sequence of steps described is illustrative only and can be varied without departing from the disclosure. Thus, unless otherwise stated the steps described are so unordered meaning that, when possible, the steps can be performed in any convenient or desirable order.

[0126] The disclosure should not be seen in any way restricted to the embodiments described and a person with ordinary skill in the art will foresee many possibilities to modifications thereof.

[0127] The above described embodiments are combinable. The following claims further set out particular embodiments of the disclosure.