Abstract
The present disclosure provides for the use of a temperature controlled shipping container as a method for the packaging and distribution of mammalian cells, e.g., in induced pluripotent stems cells (iPSc), and various organ cells differentiated from the iPSc. In one embodiment, a shipping box that utilizes a vacuum, desiccant and insulation construct is employed to package iPSc and their derivatives or progeny that are or have been deposited onto microwell plates or tubes that contain a nutrient solution during storage and shipping. The present system and method can maintain the cells in the containment vessels at or below 10 C. for at least 24 hours and in one embodiment up to 96 hours.
Claims
1. A method of preparing living cells for packaging and distribution, comprising: providing a receptacle having a compartment, wherein at least a portion of the compartment comprises an insulating material having a predefined thickness and predefined thermal conductivity which can maintain a temperature of about 1 C. to about 10 C. for at least 24 hours in the compartment; providing a sorption device which comprises a passive heat absorbing material; providing at least one cell culture substrate having living cells in media; activating the sorption device; placing the cell culture substrate and the sorption-cooling device in proximity in the compartment of the receptacle; and sealing the receptacle to allow for a temperature of about 1 C. to about 10 C. for at least 24 hours in the compartment.
2. The method of claim 1 wherein the cells are iPSc.
3. The method of claim 1 wherein the cells are differentiated iPSc.
4. The method of claim 1 wherein the insulating material can maintain a temperature of about 4 C. to about 10 C. for at least 24 hours.
5. The method of claim 4 wherein the temperature is maintained for at least 96 hours.
6. The method of claim 1 wherein the substrate is a multiwell plate.
7. The method of claim 1 further comprising covering the substrate with a lid so as to provide a water-proof barrier.
8. The method of claim 1 wherein the compartment in the receptacle has a temperature of about 4 C. to about 10 C. 24 hours after sealing.
9. The method of claim 1 wherein the compartment further comprises a temperature sensor.
10. The method of claim 1 wherein the cells have a viability of at least 25% up to at least 90% 24 hours after sealing.
11. The method of claim 1 wherein the cells have a viability of at least 25% up to at least 90% 96 hours after sealing.
12. The method of claim 2 wherein at least 25% up to at least 90% of the cells are capable of differentiation 24 to 96 hours after sealing.
13. The method of claim 1 wherein the sorption device absorbs heat so that the temperature in the compartment is about 1 C. to about 10 C.
14. The method of claim 1 wherein the volume of the compartment provides for adequate oxygen during distribution.
15. The method of claim 14 wherein a plurality of cell culture substrates are placed in the compartment.
16. The method of claim 15 wherein different types of cells are in each substrate.
17. The method of claim 1 wherein the seal provides a water-proof barrier.
18. The method of claim 1 wherein the seal provides a barrier that prevents or inhibits oxygen exchange.
Description
BRIEF DESCRIPTION OF THE DRAWINGS
[0011] FIG. 1 shows an example of a sorption box utilized in the present disclosure.
[0012] FIG. 2 shows an interior view of an exemplary sorption box of the present disclosure.
[0013] FIG. 3 shows an exemplary sorption module used in combination with the insulated box to enable hypothermic cooling.
[0014] FIG. 4 shows water loaded petri dishes used to simulate a biological load to be cooled.
[0015] FIG. 5 shows the cooling curves of the control thermometer outside the box and the thermometer temperature inside the storage compartment of the box 14 hours after activation.
[0016] FIG. 6 shows the temperature profile of a shipping box exposed to extreme outside temperature vs. ambient conditions.
[0017] FIG. 7 shows the temperature profile after 48 hours of activation.
[0018] FIG. 8 shows the temperature profile after 70 hours of activation.
[0019] FIG. 9 shows a cross sectional view of the cells loaded into a 384 microplate of the present disclosure that is loaded into the cooling chamber of the sorption box.
DETAILED DESCRIPTION
[0020] The present disclosure provides for storage and shipping of living cells, in particular iPSc and cells differentiated from the iPSc, by the use of a package that does not use any ice, or cryogenic materials such as liquid nitrogen or frozen carbon dioxide, in a very small, easy to ship form for extended periods of time.
[0021] Referring to FIG. 1, this shows one example of a shipping device 10 useful in the present disclosure. It is a sorption-cooling device that can keep an internal insulated storage compartment contained within at a controlled temperature for extended periods of time. This device is described in U.S. Pat. Nos. 6,559,797, 6,688,132, 6,701,724, and 6,968,711, the disclosures of which are incorporated by reference herein. Other passive cooling devices, and other shipping devices that include insulating material, may be employed in the methods without departing from the scope of the disclosure. The details of the device theory and operation are described in detail in the aforementioned art and it is assumed that one skilled in the art would understand its operation and performance characteristics. In the present disclosure, the device is employed in packaging and shipping live mammalian cells, e.g., human iPSc and differentiated cells from the iPSc such as heart, brain, liver, kidney, pancreatic, connective tissue, skin, cartilage and blood cells to name a few.
[0022] FIG. 2 shows the interior of the cooling package useful in the present disclosure. The exterior is a cardboard shell into which in the presence of insulation 20 forms an insulated compartment 30. The insulation 20 may be comprised of open and/or closed cell foam materials that are either formed of organic materials such as polymers or inorganic materials such as aerogels. The foam thickness and thermal conductivity determine the ultimate time that a biological material can be stored at hypothermic conditions.
[0023] FIG. 3 show the sorption device 40 of the present disclosure, which is responsible for removing the heat in the storage compartment. The sorption device is a closed system where a liquid such as water is evaporated under vacuum and absorbed by a desiccant in the upper part of the chamber. This passive cooling engine can be sized accordingly to enable longer or shorter hypothermic cooling times based on the loading in the cooling chamber.
[0024] Referring to FIG. 4, this is an example of a hypothermic simulation of a biological loading into the cooling chamber 30 of the present disclosure. Into the cooling chamber 30 was loaded 8 petri dishes 50 each with 5 milliliters of water to mimic a biological sample. Also included into the cooling chamber was a wireless Bluetooth temperature sensor 55 to monitor the internal temperature with time. The sorption device 40 was activated and then positioned over the cooling chamber.
[0025] FIG. 5 shows a comparison of the ambient outside temperature on the left image vs. the inside temperature right hand image of the cooling device 77 minutes after activation of the sorption chamber. The outside temperature is 77.65 F. and the inside cooling chamber is 36.70 F. At the onset it appears that the sorption-cooling device can establish and maintain the correct hypothermic temperature of biological samples.
[0026] FIG. 6 shows a comparison of the shipping box after 18 hours at room temperature (ambient conditions outside of the package) and then exposed to an outside temperature of 96.39 F. for 6 hours. The top image shows the outside temperature and the bottom image the cooling chamber temperature under the same conditions. As can be seen from the data even though the ambient conditions were extreme the cooling chamber where the biologic simulation was being monitored only rose to 38.91 F. well below hypothermic conditions that is generally considered to be 50 F. The total duration of the test at this point was 24 hours.
[0027] FIG. 7 shows the data after the biologic simulation for 48 hours. As you can see from the data that the ambient temperature has averaged 77 F. top image while the bottom image shows the internal temperature was maintained hypothermic at 36.16 F. during the period from 24 to 48 hours of testing.
[0028] FIG. 8 shows the test results after 70 hours. The top image showing the outside ambient conditions at 77.79 F. and the internal final storage temperature after 70 hours of testing rising to 41.61 F. The testing was stopped after 70 hours because it was determined that the sorption cooling device was adequate for shipping cells, such as iPSc and differentiated cells, anywhere in the world by using express carriers.
[0029] FIG. 9 shows a typical biologic microplate that can be utilized with the present disclosure. It is generally an nn array of wells and in one embodiment 34 in total. The well plate 70 is generally molded of typical thermoplastic polymers. The wells 80 contain biological material such as cells and in one embodiment iPSc or cells differentiated into various human organ cells. A lid 90 is placed over the wells to contain the cells and liquid nutrient media from spilling during shipping and handling. The lid sealing material is in one embodiment a water barrier but allows for the exchange of gasses such as oxygen and carbon dioxide so that the living cells can carry on respiration during hypothermic shipping and handling.
[0030] The above discussion is meant to be illustrative of the principles and various embodiments of the present disclosure. Numerous variations and modifications will become apparent to those skilled in the art once the above disclosure is fully appreciated. It is intended that the following claims be interpreted to embrace all such variations and modifications.
