DEVICES AND METHODS FOR THAWING BIOLOGICAL MATERIAL

Provided herein are devices and methods for thawing frozen biological material.

Provided herein are devices and methods for thawing frozen biological material.

Frozen biological material (e.g., cells) is regularly used in both laboratory and clinical settings. However, upon thawing such biological material, it is often difficult to obtain consistent results (e.g., in terms of cell viability, etc.) due to varying and non-ideal conditions under which the biological material is thawed. Therefore, there remains a need for devices which can safely and rapidly thaw biological material in a mariner that maintains the optimal condition of the biological material.

In one embodiment, provided herein is an improved thawing apparatus, wherein said apparatus comprises one or more compartments into which a container comprising frozen biological material may be thawed, wherein said one or more compartments comprise compressible material (e.g., compressible gasket material). Such thawing apparatuses are advantageous in that they can accommodate biological material containers (e.g., tubes or vials) of varying sizes, and also ensure that the containers (e.g., tubes or vials) that contain the biological material to be thawed are thoroughly in contact with the heat source. In certain embodiments, the improved thawing apparatuses comprising compressible material described herein are similar in design to dry block thawing apparatuses known in the art, but possess the advantage of comprising compressible material.

In another embodiment, provided herein is an improved thawing apparatus, wherein said apparatus comprises one or more compartments into which a container comprising frozen biological material may be thawed, wherein said one or more compartments comprises a cuff which can be placed around containers (e.g., tubes or vials) of varying size and manipulated so that it comes into thorough contact with the container. Such cuffs can be filled with a heated substance (e.g., heated fluid), the levels of which can be adjusted (e.g., filled with more or less fluid) to ensure accommodation of containers of varying size and to ensure that thorough contact between the container and the heating source. Alternatively, such cuffs may comprise a heated material which can be expanded by the application of pressure (e.g., air pressure; fluid pressure, e.g., oil pressure, water pressure, or the like) to ensure accommodation of containers of varying size and to ensure that thorough contact between the container and the heating source. In certain embodiments, the improved thawing apparatuses comprising cuffs described herein are similar in design to dry block thawing apparatuses known in the art, but possess the advantage of comprising cuffs, the size of which can be modified/adjusted.

In another embodiment, provided herein is an improved thawing apparatus, wherein said apparatus comprises (i) a sealed compartment comprising material that undergoes a phase change when heated to a certain temperature (e.g., paraffin or beeswax), (ii) a compartment into which a source of heat (e.g., boiling water) can be added, and (iii) at least one compartment into which a container (e.g., tube or vial) containing biological material can be placed. The improved thawing apparatus functions as follows: (i) the phase change of the phase change material is effected by addition of a heat source to the appropriate compartment of the apparatus, e.g., by adding boiling water to the compartment of the apparatus; (ii) once the phase change material has reached its melting point, the container(s) (e.g., tube or vial) containing the biological material to be thawed is then placed into the appropriate compartment of the apparatus; and (iii) as the frozen material cools the phase change material, the phase change material will remain at is melting point, which is suitable to result in the thawing of the biological material.

In another embodiment, provided herein is an improved thawing apparatus, wherein said apparatus comprises (i) a sealed compartment comprising a chemical material (e.g., iron) that heats to a certain temperature following exposure to a given condition (e.g., exposure to oxygen) and (ii) at least one compartment into which a container (e.g., tube or vial) containing biological material can be placed. The improved thawing apparatus functions as follows: (i) the apparatus is heated by exposing the chemical material to a given condition (e.g., oxygen); and (ii) once the apparatus is at its desired temperature, the container(s) (e.g., tube or vial) containing the biological material to be thawed is then placed into the appropriate compartment of the apparatus; resulting in the thawing of the biological material.

4. BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1is a perspective view of an exemplary thawing apparatus of the present invention having compressible material disposed in the compartments.

FIGS. 2 and 3are perspective sectional views of the thawing apparatus ofFIG. 1taken along the view line F2, wherein the apparatus illustrated inFIG. 3has a container inserted in a compartment.

FIG. 4is a perspective view of an exemplary thawing apparatus of the present invention having cuffs disposed in the compartments.

FIGS. 5 and 6are perspective sectional views of the thawing apparatus ofFIG. 4, taken along the view line F5, wherein the apparatus illustrated inFIG. 6has a container inserted in a compartment.

FIGS. 7 and 8are perspective views of an exemplary thawing apparatus of the present invention having cuffs, wherein the container is disposed outside the cuffs in a contracted or deflated condition inFIG. 7and the container is disposed inside the cuffs in an expanded or inflated condition inFIG. 8.

FIG. 9is a perspective view of an exemplary thawing apparatus of the present invention having a phase change material.

FIG. 10is a perspective sectional view of the thawing apparatus ofFIG. 9taken along the view line F10.

FIG. 11is a perspective sectional view of alternative thawing apparatus taken along the view line F10.

FIG. 12is a perspective view of an exemplary thawing apparatus of the present invention having a chemical material.

FIG. 13is a perspective sectional view of the thawing apparatus ofFIG. 12taken along the view line F13.

FIG. 14is a perspective sectional view of an alternative thawing apparatus taken along the view line F13and having controllable mechanisms.

FIG. 15is a perspective sectional view of an alternative thawing apparatus taken along the view tine F13and having controllable mechanisms and a controller operatively coupled to an external computer.

5. DETAILED DESCRIPTION

Provided herein are improved devices for thawing frozen biological material (e.g., cells). The devices described herein are designed to ensure that the container (e.g., tube or vial) which holds the frozen biological material to be thawed is consistently and thoroughly in contact with the source of heat and thus promotes rapid thawing of the biological material. The devices described herein comprise controllable temperature settings, so that they may be utilized at any thawing temperature desired by the individual using the device (e.g., 23° C., 37° C., 50° C., 100° C., etc.), and also so that the temperature can be increased or decreased during the thawing process (e.g., the temperature can be increased or decreased in 1, 2, 3, 4, 5 or more degree intervals by, e.g., pressing a button). In certain embodiments, the devices described herein additionally comprise one or more components that enable agitation (e.g., gentle shaking or rotation) of the biological material to be thawed during the thawing process.

Dry block thawing apparatuses currently known in the art are problematic in that the compartments into which the biological material containers are placed are static, i.e., they are generally made of solid metal and are thus not able to accommodate containers (e.g., tubes or vials) of varying sizes. This design leads to various problems, including containers e.g., tubes or vials) becoming stuck in the compartments into which they are placed (due to the tubes being too large for the compartment) or containers (e.g., tubes or vials) loosely fitting into the compartments they are placed (due to the tubes being too small for the compartment). Moreover, dry block thawing apparatuses currently known in the art do not promote ideal contact of the biological material containers with the source of heat, and this issue is worsened in cases where the containers loosely fit in the compartments of the dry block thawing apparatus.

Water bath thawing apparatuses currently known in the art also are problematic for various reasons. First, water bath thawing apparatuses typically require that the container that contains the biological material to be thawed is often not exposed to an ideal amount of the heat source, similar to the situation with known dry block thawing apparatuses. Further, water bath thawing apparatuses known in the art are very likely to become contaminated, thus introducing the risk that the biological material to be thawed also will become contaminated.

The improved thawing apparatuses described herein overcome the issues that occur with known dry block thawing apparatuses and water bath thawing apparatuses, as well as other thawing apparatuses known in the art by, without limitation: (i) promoting significantly greater amount of contact between the container (e.g., tube or vial) that contains the biological material than that which occurs with thawing apparatuses currently known in the art; and (ii) being able to accommodate containers of various sizes.

Referring toFIGS. 1 to 3, an exemplary thawing apparatus having compressible material, e.g., compressible material with a high heat transfer coefficient or high heat conduction, is provided in accordance with the principles of the present invention. Thawing apparatus10includes a plurality of compartments11having compressible material12disposed therein, and heating element13. Each compartment11may be configured to receive container20comprising frozen biological material21. Compressible material12(e.g., compressible gasket material) is configured to line compartment11as illustrated inFIGS. 1 and 2and to compress when container20is inserted in compartment11as illustrated inFIG. 3. Heating element13may be a conventional heating element and is configured to heat thawing apparatus10to a temperature suitable to thaw frozen biological material21disposed in container20. Thawing apparatus10may advantageously accommodate one or more biological material containers (e.g., tubes or vials) of varying sizes, and also ensure that the containers (e.g., tubes or vials) having the biological material to be thawed are thoroughly in contact with the heat from apparatus10. In certain embodiments, the improved thawing apparatuses comprising compressible material described herein are similar in design to dry block thawing apparatuses known in the art, but possess the advantage of comprising compressible material. In certain embodiments, the compressible material is a known compressible gasket material. In a specific embodiment, the compressible material is silicone, rubber, or sponge (or a combination thereof).

Referring now toFIGS. 4 to 6, an alternative exemplary thawing apparatus having cuffs is provided in accordance with the principles of the present invention. Thawing apparatus40includes a plurality of compartments41each having cuff42disposed therein, heating element43, pump fluid44, pump45, and channels46. Each compartment41may be configured to receive container50comprising frozen biological material51. Cuff42is configured to line compartment41as illustrated inFIGS. 4 and 5and to expand when container50is inserted in compartment41as illustrated inFIG. 6by pumping pump fluid44using pump45into cuff42through respective channel46. Heating element43may be a conventional heating element disposed in or near pump fluid44and is configured to heat pump fluid44to a temperature suitable to thaw frozen biological material51disposed in container50. Alternatively, heating element43may be disposed outside of pump fluid44and configured to heat apparatus40to thaw frozen biological material51. Pump45may be a conventional pump and is configured to pump pump fluid44to and from a respective cuff42using a respective channel46to cause the cuff to expand or contract. As will be apparent to one of ordinary skill in the art, pump45and/or channels46may include one or more valves suitable for controlling flow of pump fluid44to or from cuffs42. Containers (e.g., tubes or vials) of varying size may be placed in compartments41and cuffs42may be manipulated to come into contact with the container. In one embodiment, such cuffs may comprise a heated material which may be expanded by the application of pressure (e.g., air pressure; fluid pressure, e.g., oil pressure, water pressure, or the like) to ensure accommodation of containers of varying size and to ensure that thorough contact between the container and the heating source. In certain embodiments, the improved thawing apparatuses comprising cuffs described herein are similar in design to dry block heating/warming apparatuses known in the art, but possess the advantage of comprising cuffs, the size of which can be modified/adjusted.

Referring now toFIGS. 7 to 8, another alternative exemplary thawing apparatus having cuffs is provided in accordance with the principles of the present invention. Thawing apparatus70includes at least one cuff71, connectors72, fluid source pump73, and at least one tube74. Cuff71is configured to receive container80comprising frozen biological material81. As will be apparent to one of ordinary skill in the art, more than one cuff71may he used as illustrated and connected via connectors72or one cuff shaped and sized to receive container80may be used. Fluid source pump73is configured to store fluid and may include a conventional heating element for heating the fluid and a conventional pump for pumping the heated fluid to cuff(s)71through tube74and optionally to more than one cuff71through connectors72. Cuff71is configured to contract when fluid is pumped therefrom as illustrated inFIG. 7and to expand when fluid is pumped therein as illustrated inFIG. 8. Cuffs71may be placed around containers (e.g., tubes or vials) of varying size and manipulated to come into contact with the container. In one embodiment, such cuffs may comprise a heated material which may be expanded by the application of pressure (e.g., air pressure) to ensure accommodation of containers of varying size and to ensure that thorough contact between the container and the heating source.

Referring now toFIGS. 9 to 10, yet another exemplary thawing apparatus is provided in accordance with the principles of the present invention. Thawing apparatus90includes housing91, a plurality of compartments92, a plurality of heating compartments93, and phase change material94. Each compartment92may be configured to receive container100comprising frozen biological material101. Housing91is configured to seal phase change material94therein. Each heating compartment93is configured to receive a heated material (e.g., boiling water) at a temperature suitable to cause phase change material94(e.g., paraffin or beeswax) to undergo a phase change. Thawing apparatus90may function as follows: (i) the phase change of phase change material94is effected by addition of a heat source to heating compartment93of apparatus90, e.g., by adding boiling water to the compartment of the apparatus; (ii) once phase change material94has reached its melting point, container(s)100(e.g., tube or vial) containing frozen biological material101to be thawed is then placed into compartment92of apparatus90; and (iii) as frozen biological material101cools phase change material94, phase change material94remains at is melting point, which is suitable to result in the thawing of the frozen biological material.

Referring toFIG. 11, thawing apparatus90′ is constructed substantially identically to thawing apparatus90ofFIGS. 9 and 10, wherein like components are identified by like-primed reference numbers. Thus, for example, housing91′ inFIG. 11corresponds to housing91ofFIGS. 9 and 10, etc. As will be observed by comparingFIGS. 10 and 11, apparatus90′ does not include heating compartment93and includes heating element95. Heating element95may be disposed in or near phase change material94′ and may be a conventional heating element configured to heat phase change material94′ to a suitable temperature to cause phase change material94′ to undergo a phase change.

Referring now toFIGS. 12 to 13, an alternative exemplary thawing apparatus having a chemical material is provided in accordance with the principles of the present invention. Thawing apparatus120includes housing121, a plurality of compartments122, reactant source pump123having reactant124, tube(s)125and chemical material126. Each compartment122may be configured to receive container130comprising frozen biological material131. Housing121is configured to seal chemical material126therein. Reactant source pump123is configured to store reactant124(e.g., oxygen) and may include a conventional pump for pumping reactant124to chemical material126(e.g., iron) through tube(s)125. Chemical material126is configured to heat to a suitable temperature to thaw frozen biological material131following exposure to a given condition (e.g., exposure to a reactant such as oxygen from reactant source pump). Thawing apparatus120may function as follows: (i) chemical material126is heated by exposing chemical material126to a given condition (e.g., reactant from reactant pump source); and (ii) once the apparatus is at its suitable temperature, container(s)130(e.g., tube or vial) containing biological material131to be thawed is then placed into the appropriate compartment122of apparatus120; resulting in the thawing of biological material131. In a specific embodiment, the chemical material is iron and the given condition is exposure to oxygen. In another specific embodiment, the chemical material is sodium acetate. In another specific embodiment, the chemical material is liquefied petroleum gas (lpg) and the given condition is exposure to platinum.

In certain embodiments, the improved thawing apparatuses described herein result in a 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, or greater than 50% increase in contact between the container (e.g., tube or vial) that contains the biological material than that which occurs with a thawing apparatus currently known in the art (e.g., a standard dry block thawing apparatus or a standard water bath thawing apparatus).

Referring toFIGS. 14 and 15, thawing apparatuses120′ and120″ are constructed substantially identically to thawing apparatus120ofFIGS. 12 and 13, wherein like components are identified by like-primed reference numbers. Thus, for example, housing121′ inFIG. 14and housing121″ inFIG. 15correspond to housing121ofFIGS. 12 and 13, etc. As will be observed by comparingFIGS. 13 and 14, apparatus120′ further includes one or more controllable mechanisms140coupled by at least one shaft142to motor143. Controllable mechanisms140are configured to agitate container(s)130(e.g., tube or vial) containing biological material131to be thawed. Motor143may be a conventional motor configured to agitate controllable mechanism140by moving shaft(s)142. In one embodiment, motor143is configured to agitate controllable mechanism140so as to shake container(s)130(e.g., tube or vial) containing biological material131to be thawed, e.g., shakes the container up and down or left to right as illustrated, or both. In another embodiment, motor143is configured to agitate controllable mechanism140to rotate container(s)130(e.g., tube or vial) containing biological material131to be thawed, e.g., the controllable mechanism rotates the container in a clockwise or counter-clockwise manner.

As will be observed by comparingFIGS. 14 and 15, apparatus120″ further includes controller150for controlling components of thawing apparatus120″ including motor143′. Controller150may include any one or more microprocessors, controllers, digital signal processors (DSPs), application specific integrated circuits (ASICs), field-programmable gate arrays (FPGAs), or equivalent discrete or integrated digital or analog logic circuitry, and the functions attributed to controller150herein may be embodied as software, firmware, hardware, or any combination thereof. Controller150may include a memory for storing data related to use of thawing apparatus120″, such as a selected program for agitating container(s). The memory may store program instructions that, when executed by controller150, cause controller150and apparatus120″ to provide the functionality ascribed to them herein. The memory of controller150also may store software downloaded thereon or implemented as a program product and stored on a tangible storage device such as machine-readable medium, e.g., tape, compact disk (CD), digital versatile disk (DVD), blu-ray disk (BD), flash drive, and so forth, external nonvolatile memory device, cloud storage, or other tangible storage medium. The software may include computer executable instructions for controlling apparatus120″. Optionally, controller150may include suitable components for wired and/or wireless communication with external computer151, illustratively a laptop computer. External computer151may have software downloaded thereon for controlling apparatus120″. In one embodiment, the software on external computer151allows a practitioner (e.g., a lap technician) to select a predefined program and/or create a program for agitating container(s). As will be readily apparent, thawing apparatus120″ and external computer151are not to scale.

In certain embodiments the apparatuses described herein comprise controllable temperature settings, so that they may be utilized at any thawing temperature desired by the individual using the device (e.g., 23° C., 37° C., 50° C., 100° C., etc.), and also so that the temperature can be increased or decreased during the thawing process (e.g., the temperature can be increased or decreased in 1, 2, 3, 4, 5 or more degree intervals by, e.g., pressing a button). In certain embodiments, the apparatuses described herein can be programmed to thaw at a specific temperature (e.g., 42° C.) for a specific period of time, followed by a shift to a different temperature (e.g., 37° C.) for a specific period of time.

In certain embodiments, the apparatuses described herein comprise one or more methods for alerting the practitioner that the thawing cycle of a certain biological material has been completed, e.g., the apparatus may comprise a bell, a buzzer, a flashing light or any combination thereof, or any other similar alerting mechanism. In certain embodiments, the device may comprise instructions for thawing certain frozen biological materials that are frozen in a specific volume, and also may provide times for which said frozen biological materials that are frozen in a specific volume will be thawed using the apparatus.

The duration of time for thawing of a biological material using a thawing apparatus described herein will depend on a variety of factors including, without limitation, the volume of the frozen biological material, the thawing apparatus used, whether or not the practitioner wishes to agitate (e.g., rotate or shake) the container containing the biological material, and the temperature the practitioner desires to thaw at. Those of skill in the art will appreciate how the improved thawing apparatuses can be adjusted to accommodate the desired timing and degree of thawing.

As will be apparent to one of ordinary skill in the art, components of the thawing apparatuses described herein may be used separately or in combination with one another. For example, controllable mechanism(s)140′, shaft(s)142′, motor143′, and/or controller150ofFIG. 15may be incorporated in thawing apparatus10, thawing apparatus40, thawing apparatus70, and/or thawing apparatus90, etc. without departing from the principles of the present invention.

Any container suitable for freezing biological material can be used in the thawing apparatuses described herein. In certain embodiments, the containers comprise tubes. In certain embodiments, the containers comprise vials. In certain embodiments, the containers comprise bags. Such containers may be made of any material known in the art, e.g., glass, plastic, polystyrene, etc.

The containers be used in the thawing apparatuses described herein may be of any size known in the art. In certain embodiments, the containers have a volume of 100 μl to 500 μl, 500 μl to 1 ml, 1 ml to 2 ml, 1 ml to 5 ml, 5 ml to 10 ml, 10 ml to 25 ml, 25 ml to 50 ml, 50 ml to 100 ml, or 100 ml to 1 L. In certain embodiments the containers have a volume of more than 1 L.

5.3 Biological Material

Any biological material can be thawed in a container using the thawing apparatuses described herein. In a specific embodiment, the biological material comprises cells, e.g., cryopreserved cells. In another specific embodiment, the biological material comprises intact tissue or organ. In another specific embodiment, the biological material comprises blood, e.g., human or animal blood.

The devices and methods disclosed herein are not to he limited in scope h the specific embodiments described herein. Indeed, various modifications of the devices and methods in addition to those described will become apparent to those of skill in the art from the foregoing description and accompanying figures. Such modifications arc intended to fall within the scope of the appended claims.