IP Library › Granted Patent US 12,195,748
Granted Patent B2
US 12,195,748 · App. 17/337,177 · Granted Jan 14, 2025

Delivery across cell plasma membranes

Inventors: Michael Maguire (Dublin, IE); Shirley O'Dea (Maynooth, IE)
Assignee: Avectas Limited
C12N15/895A61K31/473A61K31/7115A61K31/721A61P43/00B05B1/02B05B7/2491C12M25/14C12M33/04C12M35/04C12N5/00C12N5/0018C12N5/0636C12N5/0656C12N5/0662C12N15/113C12N15/87C12N2310/14C12N2500/12C12N2500/30C12N2500/34C12N2500/60
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Quick Facts
Patent No.
US 12,195,748
App. No.
17/337,177
Granted
Jan 14, 2025
Kind
B2
Abstract

Delivering a payload across a plasma membrane of a cell includes providing a population of cells and contacting the population of cells with a volume of an aqueous solution. The aqueous solution includes the payload and alcohol content greater than 5 percent concentration. The volume of the aqueous solution may be a function of exposed surface area of the population of cells, or may be a function of a number of cells in the population of cells. Related compositions, apparatus, systems, techniques, and articles are also described.

Claims (34)

1. An apparatus for delivering a payload across a plasma membrane of a cell, the apparatus comprising:

a pneumatic generator configured to produce a gas under a pressure;

an atomizer operatively coupled to the pneumatic generator, wherein the atomizer is oriented to contact a population of cells with an aqueous solution;

a valve between the pneumatic generator and atomizer, the valve switchable between a closed position for preventing the gas under the pressure from activating the atomizer and an open position for allowing the gas under the pressure to activate the atomizer to produce colloidal droplets from the aqueous solution; and

a reservoir containing a volume of the aqueous solution, the aqueous solution including the payload and an alcohol at greater than 2 percent concentration (v/v), less than 46 mM salt, less than 121 mM sugar, and less than 19 mM buffering agent, wherein the salt is present in the aqueous solution and selected from a group consisting of NaCl, KCl, Na 2 HPO 4 , C 2 H 3 O 2 NH 4 , and KH 2 PO 4 .

2. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets having a diameter of 500 nm to 100 micrometers.

3. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets having a diameter of between 1 and 30 micrometers.

4. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets that spray at a distance of 21 millimeter (mm) to 41 mm from the population of cells.

5. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets at a distance of less than 31 millimeter (mm) from the population of cells.

6. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets at a location that is a distance from the population of cells and wherein the pressure is 0.05 bar per millimeter distance between the population of cells and the location.

7. The apparatus of claim 1 , wherein the atomizer is arranged to produce colloidal droplets at a location that is a distance from the population of cells, and wherein the distance is approximately twice a diameter of an area of the population of cells.

8. The apparatus of claim 1 , wherein the apparatus is configured to produce colloidal droplets having a total volume between 6.0×10 −7 microliter per cell and 7.4×10 −4 microliter per cell.

9. The apparatus of claim 1 , wherein the volume is between 9.3×10 −6 microliter per cell and 2.8×10 −5 microliter per cell.

10. The apparatus of claim 1 , wherein the volume is about 1.9×10 −5 microliters per cell, wherein about is within 10 percent.

11. The apparatus of claim 1 , wherein the volume is between 2.6×10 −9 microliter per square micrometer of exposed surface area and 1.1×10 −6 microliter per square micrometer of exposed surface area.

12. The apparatus of claim 1 , wherein the volume is between 5.3×10 −8 microliter per square micrometer of exposed surface area and 1.6×10 −7 microliter per square micrometer of exposed surface area.

13. The apparatus of claim 1 , wherein the volume is about 1.1×10 −7 microliter per square micrometer of exposed surface area, wherein about is within 10 percent.

14. The apparatus of claim 1 , wherein a total volume of aqueous solution of 20 μl is delivered to a cell-occupied area of about 1.9 cm 2 , wherein about is within 10 percent.

15. The apparatus of claim 1 , wherein a total volume of aqueous solution of 10 μl is delivered to a cell-occupied area of about 0.95 cm 2 , wherein about is within 10 percent.

16. The apparatus of claim 1 , wherein the alcohol is selected from methanol, ethanol, isopropyl alcohol, butanol and benzyl alcohol.

17. The apparatus of claim 1 , wherein the alcohol comprises ethanol at a concentration of 2 to 30%.

18. The apparatus of claim 1 , wherein the population of cells is in contact with the aqueous solution for 0.1-10 minutes prior to adding a second volume of buffer or culture medium to submerse or suspend said population of cells, and wherein the buffer or culture medium comprises phosphate buffered saline (PBS).

19. The apparatus of claim 1 , wherein the valve has a switching speed of less than 200 milliseconds.

20. The apparatus of claim 1 , wherein the valve has a switching speed of less than 50 milliseconds.

21. The apparatus of claim 1 , wherein the valve has a switching speed of about 50 milliseconds, wherein about is within 10 percent.

22. The apparatus of claim 1 , wherein the valve has a switching speed of between 50 and 200 milliseconds.

23. The apparatus of claim 1 , wherein said gas comprises nitrogen, ambient air, or an inert gas.

24. The apparatus of claim 1 , wherein the pressure is 1.0 to 2.5 bar.

25. The apparatus of claim 1 , wherein the atomizer, when the valve is in the open position, produces colloidal droplets having a diameter less than 100 micrometers.

26. The apparatus of claim 1 , wherein the atomizer, when the valve is in the open position, produces colloidal droplets having a diameter less than 30 micrometers.

27. The apparatus of claim 1 , wherein greater than 80 percent of the colloidal droplets produced by the atomizer have the diameter that is less than 30 micrometers.

28. The apparatus of claim 1 , wherein greater than 99 percent of the colloidal droplets produced by the atomizer have the diameter that is less than 30 micrometers.

29. The apparatus of claim 1 , wherein the atomizer, when the valve is in the open position, produces colloidal droplets having a diameter between 30 and 100 micrometers.

30. The apparatus of claim 1 , wherein the atomizer, when the valve is in the open position, produces colloidal droplets having a diameter between 30 and 50 micrometers.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Feb 14, 2023
From: MAGUIRE, MICHAEL; O'DEA, SHIRLEY
To: PROFECTOR LIFE SCIENCES LIMITED
Reel/Frame 062683/0616 →
CHANGE OF NAME Recorded Feb 14, 2023
From: PROFECTOR LIFE SCIENCES LIMITED
To: AVECTAS LIMITED
Reel/Frame 062746/0935 →
Priority Claims (3)
GB 1419011 · Oct 24, 2014 · national
GB 1419012 · Oct 24, 2014 · national
GB 1419013 · Oct 24, 2014 · national
Continuity (4)
Continuation 17092161 · Nov 6, 2020
Continuation 16841236 · Apr 6, 2020
Continuation 15521192
Related Publication 20210324419A1 · Oct 21, 2021
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