Cell growth device and uses thereof

Abstract

A cell growth device is provided, comprising: (a) a device for cell storage or cell culture; and (b) a means or unit for delivering an air flow to the device for cell storage or cell culture. Also provided is an in vitro method of inducing cell growth by placing a plurality of cells in the cell growth device described herein.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to Australian Provisional Application No. 2022900445, filed on 25 February 2022, the contents of which are incorporated herein by reference in their entirety.

[0002] The present invention relates to a cell growth device. Methods of growing cells in vitro using the cell growth device disclosed herein are also provided. [Background technology]

[0003] Cell culture refers to the process that allows cells obtained from animals, humans and plants to grow outside of their natural environment. Cell culture using various cell culture devices is central to experimental cell biology and cell-based therapies targeting multiple systems. The interaction between cell culture parameters and the microenvironmental conditions within the cell culture device can significantly affect the number of cell harvests and the in vivo performance of the cultured cells.

[0004] Commercial and research cell cultures are increasingly being produced using cell culture equipment such as cell culture bags, which reduce the risk of contamination and facilitate scale-up and automation. However, the full potential and effectiveness of cell culture equipment for large-scale production of cells has yet to be optimized, especially for cell therapy applications.

[0005] There is a need for more efficient cell expansion devices that allow for economical and consistent production of high quality cells on a large scale to meet Good Manufacturing Practice (GMP) regulations for Advanced Medical Products (ATMPs), such as autologous and allogeneic cell therapies, with or without genetic modification. The present invention addresses this and other needs. Summary of the Invention

[0006] In one embodiment, the present invention provides a cell growth device comprising: (a) an apparatus for cell storage or cell culture; and (b) a means or unit for delivering an air flow to the apparatus for cell storage or cell culture, the air flow being directed from the means or unit directly to a surface of the apparatus on which more than 50% of the cells are located (hereinafter the surface of the apparatus). The cell growth rate increases with the speed of the air flow compared to a cell growth device without the means or unit for delivering the air flow.

[0007] In an exemplary embodiment, the angle between the means for delivering the airflow and the surface of the device on which more than 50% of the cells rest (see * in FIG. 8A, hereafter fan angle) is greater than 45° to about 90°. In another exemplary embodiment, the % of the surface of the device (on which more than 50% of the cells rest) that is blown by or in contact with the wind or airflow is at least 50%.

[0008] Some embodiments provide an in vitro method of inducing cell proliferation by placing a plurality of cells in a cell storage or cell culture device of the cell proliferation device described herein.

[0009] The terms "invention," "the invention," "this invention," and "the present invention" used in this patent are intended to broadly refer to the subject matter of this patent and all of the appended claims. Statements containing these terms should be understood to neither limit the subject matter described herein nor limit the meaning or scope of the appended claims. The embodiments of the invention encompassed by this patent are defined by the appended claims, not this summary. This summary is a high-level overview of various aspects of the invention and introduces some of the concepts that are further described in the Detailed Description section below. This summary is not intended to identify key or essential features of the claimed subject matter, nor is it intended to be used alone to determine the scope of the claimed subject matter. The subject matter should be understood by reference to appropriate portions of the entire specification, any or all drawings, and each claim.

[0010] The invention will become more apparent from the accompanying drawings and the following detailed description. Exemplary embodiments of the present invention are described in detail below with reference to the following figures: [Brief description of the drawings]

[0011] [Figure 1] An exemplary embodiment of the invention is shown, comprising an apparatus for cell culture or cell preservation (A), a means or unit for delivering air flow to the apparatus (B) and a housing (C). The white arrow indicates the direction of the air flow and the black arrow indicates the surface of the apparatus for cell preservation or cell culture on which more than 50% of the cells are present. [Figure 2A] FIG. 1 is a line graph showing cell number and fold increase of Jurkat cells cultured in a cell growth device of the present invention (airflow speed 0.82 m / s, with fan) and in a cell culture device without a fan or airflow (no fan) (*p<0.05, **p<0.01, and ***p<0.001). [Figure 2B] 1 is a bar graph showing cell number and fold increase of Jurkat cells cultured in a cell growth device of the present invention (airflow speed 0.82 m / s, with fan) and in a cell culture device without a fan or airflow (no fan) (*p<0.05, **p<0.01, and ***p<0.001). [Figure 3A] FIG. 1 is a collection of flow cytometry analysis images showing similar CD3 expression levels of Jurkat cells cultured in a cell expansion device of the present invention (airflow speed 0.82 m / s, with fan) and in a cell culture device without a fan or airflow (fanless). [Figure 3B] FIG. 1 is a collection of flow cytometry analysis images showing the % of cell death of Jurkat cells cultured in a cell growth device of the present invention (airflow speed 0.82 m / s, with fan) and in a cell culture device without a fan or airflow (no fan). [Figure 4A] 1 is a line graph showing the effect of various airflow velocities on cell number and fold expansion of Jurkat cells cultured in a cell growth device of the present invention (delivering airflow velocities of 0.46, 0.82 and 1.19 m / s) and in a cell culture device without a fan or airflow (*p<0.05, **p<0.01, and ***p<0.001). [Figure 4B] 1 is a bar graph showing the effect of various airflow rates on cell number and fold expansion of Jurkat cells cultured in a cell growth device of the present invention (delivering airflow rates of 0.46, 0.82 and 1.19 m / s) and in a cell culture device without a fan or airflow (*p<0.05, **p<0.01, and ***p<0.001). [Diagram 5] FIG. 1 is a line graph showing cell counts of human peripheral blood mononuclear cells (PBMNC) cultured in a cell expansion device of the present invention (with a fan delivering an airflow velocity of 0.82 m / s) and in a cell culture device without a fan (fan-less). [Figure 6] FIG. 1 is a collection of photographic images showing that cell growth according to the present invention (with fan, airflow velocity 0.82 m / s) does not increase the risk of bacterial transmission through the cell culture bag. [Figure 7A] 1 is a line graph showing the cell number and the increase fold of Jurkat cells cultured in a cell proliferation device of the present invention (with fan, air flow rate of 1.19 m / s). [Figure 7B] 1 is a line graph showing the cell number and the increase fold of Jurkat cells cultured in a cell proliferation device of the present invention (with fan, air flow rate of 1.19 m / s). [Figure 7C] 7A and 7B are line graphs showing cell counts and fold increase of Jurkat cells cultured in a cell proliferation device of the present invention (with fan, air flow rate 1.19 m / s). In FIG. 7A, cells were cultured in a cell culture bag with a bag wall thickness of 0.076±0.013 mm. In FIG. 7B, cells were cultured in a cell storage bag with a bag wall thickness of 0.406±0.005 mm. In FIG. 7C, cells were cultured in a cell storage bag with a bag wall thickness of 0.473±0.127 mm. [Figure 8A] 8A is a graph showing (a) the fan angle between the direction of airflow from the fan and (b) the surface of a device for cell storage on which more than 50% of the cultured cells rest ("*" in FIG. 8A ), and the effect of the fan angle on the proliferation of Jurkat cells cultured in a cell proliferation device of the invention (having a fan delivering an airflow velocity of 1.19 m / s). [Figure 8B] 8A is a graph showing (a) the fan angle between the direction of airflow from the fan and (b) the surface of a device for cell storage on which more than 50% of the cultured cells rest ("*" in FIG. 8A ), and the effect of the fan angle on the proliferation of Jurkat cells cultured in a cell proliferation device of the invention (having a fan delivering an airflow velocity of 1.19 m / s). [Figure 8C]8A-C are graphs showing the effect of fan angle ("*" in FIG. 8A) between (a) the direction of airflow from the fan and (b) the surface of a device for cell storage on which more than 50% of the cultured cells rest, and the effect of fan angle on the proliferation of Jurkat cells cultured in a cell proliferation device of the present invention (having a fan delivering an airflow velocity of 1.19 m / s). FIG. 8A shows various fan angles. FIG. 8B and FIG. 8C are bar graphs showing the effect of various fan angles (horizontal angle (0°), vertical angle (90°), and angle 45°) on the number of cultured cells. [Figure 9] 1 is a bar graph showing the effect of surface area exposed to airflow on cultured cell number in a cell growth device of the present invention (having a fan delivering an airflow velocity of 1.19 m / s). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] As used herein, the articles "a" and "an" refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "a fan" means one fan or more than one fan.

[0013] The term "culturing" refers to the process by which cells or tissues obtained from animals, humans and plants are allowed to grow outside their natural environment.

[0014] The term "cells" includes adherent and non-adherent cells. Non-adherent cells include nucleated cells, non-nucleated cells and platelets.

[0015] All numbers herein are understood to be modified by "about." As used herein, the term "about" is meant to encompass a variation of ±10%.

[0016] Culture and propagation equipment As shown in FIG. 1, the culture and growth apparatus comprises (i) an apparatus for cell culture or cell preservation (A) and (ii) a means or unit for delivering air flow to the apparatus for cell preservation or cell culture (B), the direction of the air flow is from the means or unit directly to the surface of the apparatus on which more than 50% of the cells are present (the direction of the air flow is indicated by the white arrow, and the surface of the apparatus for cell preservation or cell culture on which more than 50% of the cells are present is indicated by the * and black arrow).

[0017] The present culture growth device is superior to similar culture platforms that deliver airflow to the equipment surface (on which more than 50% rests) but do not have the means or units to preserve the properties of the cultured cells by increasing the cell growth rate and decreasing the cell death rate.

[0018] In one embodiment, the surface of the device for cell storage or cell culture comprises more than 50%, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85%, more than 90%, more than 95%, or more than 100% of the cultured cells disposed thereon. In another embodiment, the surface of the device for cell storage or cell culture comprises about 50%-100%, about 55%-100%, about 60%-100%, about 65%-100%, about 70%-100%, about 75%-100%, about 80%-100%, about 85%-100%, about 90%-100%, or about 95%-100% of the cultured cells disposed thereon.

[0019] In some embodiments, the culture growth equipment further comprises a housing (C) as shown in Figure 1. In other embodiments, the culture growth equipment does not include a heater or dehumidifier. In other embodiments, the culture growth equipment comprises a heater or dehumidifier.

[0020] In one embodiment, a portion of the cell culture or cell storage device is gas permeable. In another embodiment, a portion of the cell culture or cell storage device has a gas permeability of 3 to 1000 barrer, or any value or range of values ​​therebetween in 1 barrer increments (e.g., about 3 barrer to 40 barrer, about 3 barrer to about 50 barrer, about 3 to 100 barrer, about 3 to 200 barrer, about 3 to 300 barrer, about 3 to 400 barrer, about 3 to 500 barrer, about 3 to 600 barrer, about 3 to 700 barrer, about 3 to 800 barrer, about 3 to 900 barrer, about 4 to 1000 barrer, about 4 to 40 barrer, about 5 to 1000 barrer, about 5 to 40 barrer).

[0021] In some embodiments, the device for cell culture is a bag, a plate, a dish, a flask, a bottle, one or more wells, or a bioreactor. In some embodiments, the device for cell culture is a device as disclosed in WO2005035728 (the contents of which are incorporated by reference), comprising a top and a bottom joined by a plurality of sides, the bottom comprising a gas permeable material, and at least a portion of the sides being constructed from a gas permeable material, such that cells can be cultured when the device is oriented in at least two positions, namely vertically and horizontally.

[0022] The thickness of the cell culture bag is about 0.01 mm to about 0.2 mm, or any value therebetween in increments of 0.01 mm, or a range of values ​​(eg, about 0.01 to about 0.19 mm, about 0.05 to about 0.15 mm, etc.).

[0023] In one embodiment of the present invention, the device for cell preservation is a cell preservation bag. The thickness of the cell preservation bag is about 0.2 mm to about 0.5 mm, or any value or range of values ​​therebetween in increments of 0.01 mm (e.g., about 0.2 mm to about 0.35 mm, about 0.2 to about 0.40 mm, about 0.2 to about 0.45 mm, about 0.2 mm to about 0.47 mm, etc.).

[0024] In another aspect of the present invention, the device for cell storage or cell culture comprises a basal medium containing appropriate supplements for cell culture. Non-limiting examples of basal medium include Basal Medium Eagle, αMEM, GMEM, DMEM, DMEM / F12, RPMI-1640, IMDM, MCDB series, modified basal medium, etc. Non-limiting examples of supplements include fetal bovine serum, fetal bovine serum, human serum, human platelet lysate, albumin, pituitary extract, tissue extract, defined or undefined chemical supplements, etc.

[0025] Non-limiting examples of means or units for delivering an air flow include a fan, a blower or an electrostatic air mover.

[0026] The distance between the means or unit for delivering the airflow and the surface of the device on which more than 50% of the cells rest is about 0.5 cm to about 35 cm, or any value or range of values ​​therebetween in 0.5 cm increments (e.g., about 1 to about 25 cm, about 1 to about 20 cm, about 1 to about 30 cm, etc.). In an exemplary embodiment, there are no objects obstructing the airflow or wind flow between the means or unit for delivering the airflow and the surface of the device.

[0027] The airflow velocity (from the device) measured at the * in FIG. 1 (the surface of the device with more than 50% of the cells on it) is 0.01 m / s to 20 m / s, or any value or range of values ​​therebetween in 0.1 cm increments (e.g., about 0.3 to 2.0 m / s, about 0.1 m / s to 2 m / s, about 0.1 m / s to 3 m / s, about 0.1 m / s to 4 m / s, about 0.1 m / s to 5 m / s, about 0.1 m / s to 15 m / s, about 0.05 m / s to 10 m / s, etc.).

[0028] 8A shows a fan angle (*), which is the angle between (a) the direction of the airflow generated by the means or unit for delivering the airflow and (b) the surface of the device for cell storage or cell culture on which more than 50% of the cultured cells are located. In one embodiment, the fan angle is greater than 45° to about 90°, about 50° to about 90°, about 55° to about 90°, about 60° to about 90°, about 65° to about 90°, about 70° to about 90°, about 75° to about 90°, about 80° to about 90°, and about 85° to about 90°.

[0029] As shown by Figure 9, the percentage of the surface of the device (with more than 50% of the cells) that is blown by or in contact with the wind or air current generated by the means or unit for delivering the air current also affects the efficiency of the cell culture. In an exemplary embodiment, the percentage of the surface of the device that is blown by or in contact with the wind or air current and has more than 50% of the cells (black arrow and * in Figure 1) is 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, 200%, 210%, 220%, 230%, 240%, 250%, 260%, 270%, 280%, 290%, 300%, 350%, 400%, 450%, 500%, 550%, 600%, 650%, 700%, 800%, 900%, 1000%, 1100%, 1200%, 1300%, 1400%, 1500%, 1600%, 1700%, 1800%, 1900%, 2100%, 2200%, 2300%, 2400%, 2500%, 2500%, 2600%, 2700%, 2800%, 2900%, 3000%, 350%, 400%, 400%, 450%, 500%, 550%, 600%, 650%, 700%, 800%, 900%, 1000%, 1100%, 1200 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% to 100%, or any range therein, for example, about 25% to 100%, about 30% ~100%, about 35%-100%, and about 40%-100%, about 45%-100%, about 50%-100%, about 55%-100%, about 60%-100%, about 65%-100%, and about 70%-100%, about 75%-100%, about 80%-100%, about 85%-100%, about 90%-100%, and about 95%-100%.

[0030] In vitro culture method for inducing cell proliferation The present invention also provides an in vitro method for inducing cell proliferation, comprising the steps of: (a) placing a plurality of cells into a cell storage or cell culture device as described herein; (b) delivering an air flow to the device in step (a) by a means or unit; The direction of airflow provides an in vitro method of directing the air from the unit directly onto the surface of the device where more than 50% of the cells reside.

[0031] In an exemplary embodiment, the angle between the means for delivering the air flow and the surface of the device on which more than 50% of the cells rest is greater than 45° to about 90°. In another exemplary embodiment, the percentage of the surface of the device (on which more than 50% of the cells rest) that is blown by or in contact with the wind or air flow also affects the efficiency of cell culture. In an exemplary embodiment, the percentage of the surface of the device (black arrow and * in FIG. 1) that is blown by or in contact with the wind or air flow and on which more than 20% of the cells rest is greater than 25% to 100%.

[0032] The in vitro methods provided herein unexpectedly increase cell proliferation rates and decrease cell death rates by placing a plurality of cells in a cell growth device with airflow directed toward the surface of the device on which more than 50% of the cells are loaded, as compared to placing a plurality of cells in a cell growth device without airflow. Furthermore, cell proliferation rates increase with increasing airflow rates.

[0033] The following examples of specific embodiments for carrying out the present invention are provided for illustrative purposes only and are not intended to limit the scope of the present invention in any way. The airflow velocities referred to in the following examples are measured at the surface of a culture bag loaded with more than 50% of the cultured cells. EXAMPLES

[0034] Example 1: Effect of airflow on cultured Jurkat cells Jurkat cells, a human T-cell leukemia cell line, were cultured at 1 × 10 6The cells were seeded into CultiLife™ 215 culture bags (commercially available from Takara, Japan) at a concentration of 1000 cells / mL and cultured in a 37°C incubator with a humidified atmosphere of 5% CO2 with or without a fan. The fan delivered an airflow velocity of 0.82 m / s. The culture medium was refreshed by adding an equal amount of the original culture medium at 2-day intervals. The final volume of the culture medium was 200 mL per bag.

[0035] FIG. 2 shows a significant increase in the number and fold increase of cultured Jurkat cells when a fan was used to provide airflow compared to when no fan was used.

[0036] Example 2: Effect of airflow on the properties and survival of cultured Jurkat cells The Jurkat cells of Example 1 cultured with or without a fan were stained with anti-human CD3-APC (1 ng / mL, BioLegend, USA) and propidium iodide (PI, 10 ng / mL). A suspension of cultured Jurkat cells was prepared and diluted to 1 × 10 with cold PBS-BSA (phosphate buffered saline containing bovine serum albumin). 7 The concentration of cells / ml was adjusted. Dispense 100 μl of cell suspension into the required number of tubes. Antibodies were added at the dilution recommended by the supplier, mixed thoroughly and incubated at 4°C for at least 30 min. Cells were washed with 1 ml of cold PBS-BSA and centrifuged at 400 g for 5 min. The supernatant was discarded and cells were resuspended in 300–500 μl of PI (propidium iodide, 10 ng / mL) / Triton X-100 (0.3%) staining solution for 15 min at 37°C or 30 min at 20°C. Tubes were transferred to ice or stored at 4°C protected from light. Cultured cells were analyzed by flow cytometry.

[0037] Cells cultured with or without the fan showed similar expression levels of CD3, suggesting that the T cell phenotype was maintained by the fan (Figure 3A). However, there was increased cell death in cells cultured without the fan (5.35%) compared to cells cultured with the fan (0.64%), as shown by PI staining. See Figure 3B.

[0038] Example 3: Effect of airflow rate on cultured Jurkat cells Jurkat cells were cultured at 1 × 10 in 50 mL RPMI-1640 containing 10% fetal bovine serum (FBS). 6 Cells were seeded into CultiLife™215 Culture Bags at a concentration of 1000 cells / mL and cultured in a 37°C incubator with a humidified atmosphere of 5% CO2 with or without a fan at various airflow rates (0.46, 0.82 and 1.19 m / s). The culture medium was refreshed at 2-day intervals by adding an equal volume to the original culture medium. The final volume of culture medium was 200 mL per bag.

[0039] 4A and 4B show that as the airflow rate increases, the cell proliferation rate increases.

[0040] Example 4: Effect of airflow on cultured human peripheral blood mononuclear cells (PBMNC) Blood samples (20 mL) were diluted with PBS at a volume ratio of 1:1, and 2 volumes of peripheral blood mononuclear cell (PBMNC) separation solution (Ficoll-paque 1.077, Cytiva) were added to a new tube. The diluted blood was gently layered on top of the Ficoll to avoid mixing together. The tube was centrifuged at 800 × g for 20-30 min. The mononuclear cells were collected in a new tube and washed twice with PBS. PBMNC were collected and cultured in the cell proliferation device of the present invention (with fan, air flow rate 1.19 m / s) and in the cell culture device without a fan in a cell storage bag (JMS, Japan) containing DuoCIK medium at 37 °C with a humidified atmosphere of 5% CO2.

[0041] FIG. 5 shows that the number of cultured human PBMNC is increased in the presence of a fan to deliver airflow to the cell culture bag, rather than in bags without a fan.

[0042] Example 5: Effect of airflow on microbial permeability of cell culture bags A cell storage bag (JMS, Japan) was filled with 200 mL of LB medium and cultured in a 37°C incubator with or without a fan to deliver an airflow velocity of 0.82 m / s. To show that the medium in the cell culture bag can support bacterial growth, 100 μL of E. coli (DH5α strain, OD 0.4) in LB medium was added to a cell culture bag containing 200 mL of LB medium. After one day of culture in a 37°C incubator, the LB medium (with bacteria added) was spread on an LB agar plate and colony formation was observed one day after culture (panel A of FIG. 6). The LB medium (without bacteria added) was collected from the cell culture bag with or without a fan and spread on an LB agar plate. No colony formation was observed after 30 days of culture, indicating that airflow into the cell culture bag does not affect the impermeability of the cell culture bag to environmental microorganisms (panels B and C of FIG. 6).

[0043] Example 6: Effect of Air Flow on Cell Culture and Cell Storage Bags 1 × 10 in 50 mL of RPMI-1640 containing 10% fetal bovine serum (FBS). 6 Jurkat cells were seeded into cell culture bag 1 (CultiLife™ 215 Culture Bag) or cell storage bags 2 and 3 (bag 2, cell storage bag commercially available from JMS, Japan; bag 3, cell storage bag commercially available from Kawasumi, Japan) at a concentration of 1.19 cells / mL. Cells were cultured in a 37°C incubator with a humidified atmosphere of 5% CO2 with or without a fan to deliver an airflow velocity of 1.19 m / s.

[0044] The thickness of the cell culture bag 1 was 0.076±0.013 mm, the thickness of the cell storage bag 2 was 0.406±0.005 mm, and the thickness of the cell storage bag 3 was 0.473±0.127 mm.

[0045] 7A to 7C show that the use of a fan increases the number of cultured cells in both the cell culture bag and the cell storage bag.

[0046] Example 7: Effect of fan angle on cell growth in cell storage bags Jurkat cells were seeded into cell storage bags according to the process of Example 1 with and without a fan to deliver an airflow velocity of 1.19 m / s at various fan angles (0°, 45° and 90°).

[0047] Figure 8B and Figure 8C show the effect of fan angle on cell proliferation. The number of cultured cells increases with increasing fan angle. Specifically, when the fan angle is greater than 45°, cell proliferation increases significantly. *, p<0.05. **, p<0.01.

[0048] Example 8: Effect of surface area permeable to or in contact with airflow on cell growth in cell storage bags Cell storage bags were seeded with Jurkat cells according to the steps of Example 1 with and without a fan to deliver an airflow velocity of 1.19 m / s.

[0049] The surface of the cell storage bag was partially covered by plastic wrap such that more than 50% of the cells were present (see * in Figure 1) and 25%, 50% and 75% of the surface were covered and not penetrated or covered by the wind or breeze generated by the fan, meaning that 75%, 50% and 25% or 100% (uncovered) of the surface of the cell storage bag was in contact with the wind or breeze generated by the fan, respectively.

[0050] FIG. 9 shows that the cell number of Jurkat cells cultured in the cell growth device of the present invention increases when the percentage of the surface permeable to wind or breeze exceeds 25%.

Claims

1. (a) a device for cell storage or cell culture; (b) a means for delivering an air flow to said device for cell storage or cell culture; A cell proliferation device comprising: A cell growth device wherein the direction of the air flow is from the means directly towards the surface of the device on which more than 50% of the cells are loaded, and the angle between the means for delivering the air flow and the surface of the device on which more than 50% of the cells are loaded is greater than 45° to about 90°.

2. The cell growth apparatus of claim 1 , wherein the device for cell storage is a cell storage bag.

3. The cell growth device of claim 1 , wherein a portion of the device for cell culture is gas permeable.

4. The cell growth device of claim 1 , wherein the device for cell culture is a bag, a plate, a dish, a flask, a tube, a well or a bioreactor.

5. The cell growth device of claim 1 , wherein the device for cell storage or cell culture comprises a culture medium.

6. 2. The cell growth apparatus of claim 1, wherein the air flow velocity is between 0.03 m / s and 20 m / s at a surface of the device on which more than 50% of the cells are loaded.

7. 10. The cell growth apparatus of claim 1, wherein the means for delivering the air flow is a fan or an electrostatic air mover.

8. 10. The cell growth apparatus of claim 1, wherein the distance between said means for delivering said air flow and said instrument is from about 1 cm to about 20 cm.

9. The cell growth apparatus of claim 1 , which does not include a heater or a dehumidifier.

10. (a) a device for cell storage or cell culture; (b) a means for delivering an air flow to said device for cell storage or cell culture; A cell proliferation device comprising: A cell growth device wherein the air flow is directed from the means directly to a surface of the device on which more than 50% of the cells are present, and greater than 25% to 100% of the surface of the device is in contact with the air flow generated by the means.

11. 1. An in vitro method for inducing cell proliferation, comprising: (a) placing a plurality of cells in the device for cell storage or cell culture of claim 1; (b) delivering, by a unit, an air flow to the device of step (a); An in vitro method in which the airflow is directed from the unit directly onto the surface of the device on which more than 50% of the cells are loaded.

12. The in vitro method of claim 11 , wherein the device for cell preservation is a cell preservation bag.

13. 12. The in vitro method of claim 11, wherein a portion of the device for cell culture is gas permeable.

14. 15. The in vitro method according to claim 14, wherein the device for cell culture is a bag, a plate, a dish, a flask, a tube, a well or a bioreactor.

15. 15. The in vitro method according to claim 14, wherein the device for cell storage or cell culture comprises a culture medium.

16. 15. The in vitro method of claim 14, wherein the air flow velocity is between 0.03 m / s and 20 m / s at the surface of the device on which more than 50% of the cells are loaded.

17. 15. The in vitro method of claim 14, wherein the unit for delivering the air flow is a fan or an electrostatic air mover.

18. The in vitro method of claim 14, wherein the distance between the unit and the device is from about 1 cm to about 20 cm.

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