Disposable cartridge for electroporation

Patent metadata

Field	Value
Publication number	US-11053470-B2
Application number	US-201615252362-A
Country	US
Kind code	B2
Filing date	Aug 31, 2016
Priority date	Sep 7, 2015
Publication date	Jul 6, 2021
Grant date	Jul 6, 2021

Abstract

The invention is directed to disposable cartridge for electroporation of cells, comprising a fluid compartment in an interior of the disposable cartridge; a first fluid port for providing cell suspension to the fluid compartment, and a second fluid port for delivering a fluid comprising at least one compound to be electroporated into the cells to the fluid compartment; a first electrode and a second electrode disposed in the fluid compartment; at least one exit port which delivers the fluid from the fluid compartment wherein the first and second fluid port have a fluid communication to a mixing channel which has a fluid communication to the fluid compartment, and a third ground electrode and a fourth ground electrode.

First claim

What is claimed is:

1. A disposable cartridge for electroporation of cells, comprising
a fluid compartment in an interior of the disposable cartridge;
a first fluid inlet port for providing a first fluid comprising a cell suspension to the fluid compartment, and a second fluid port for delivering a second fluid comprising at least one compound to be electroporated into the cells to the fluid compartment;
a first electrode and a second electrode disposed in the fluid compartment, with a voltage applied between the first and second electrodes, wherein the first and second electrodes are spaced such that an electric field of more than 2 kV/cm is formed between the first and the second electrodes;
at least one exit port which delivers the fluid from the fluid compartment, wherein the first inlet port and the second fluid ports have a fluid communication to a mixing channel which has a fluid communication to the fluid compartment; and
a third ground electrode disposed in the first fluid inlet port and a fourth ground electrode disposed in the at least one exit port, and the third and fourth ground electrodes are not between the first and the second electrodes and configured to remove charge from cell suspension, and wherein a voltage pulse is applied between the first and the second electrodes, and the third and fourth electrodes are configured to apply ground potential to the cell suspension in the first fluid inlet port and the at least one exit port.

2. The disposable cartridge of claim 1 , wherein the fluid compartment has a non-rectangular parallelogram shape with one corner higher than the others, and wherein the at least one exit port and the first fluid inlet port are disposed at the highest and lowest corners of the parallelogram fluid compartment, respectively and wherein the mixing channel has a serpentine shape and/or comprises agitator elements or a mixing chamber.

3. The disposable cartridge of claim 1 , wherein the first fluid inlet port and the second fluid port are each provided with a channel each having a different microfluidic resistance.

4. The disposable cartridge of claim 3 , wherein the first fluid inlet port and the second fluid port are each provided with a channel each having a different microfluidic resistance, so as to maintain a mixing ratio of 1:5 to 1:10 of the first fluid to the second fluid.

5. The disposable cartridge according to claim 1 , wherein the first fluid inlet port and the second fluid port are in fluid communication with at least one pump delivering fluids from the first fluid port and the second fluid into the fluid compartment.

6. The disposable cartridge according to claim 1 , wherein the at least one exit port is in fluid communication to at least one vacuum source thereby sucking fluids from the first fluid port and the second fluid into the fluid compartment.

7. The disposable cartridge according to claim 1 , wherein the mixing channel and the at least one exit port are located at opposite sides of the fluid compartment to enable bubble-free filling of the fluid compartment with fluid.

8. The disposable cartridge according to claim 1 , wherein the first electrode and the second electrode comprise titanium covered with a layer of gold.

9. A process for electroporation of cells in the disposable cartridge of claim 1 , wherein a cell suspension is provided to a first fluid inlet port as first fluid and a fluid comprising at least one compound to be electroporated into the cells is provided to the second fluid port as second fluid; the first and second fluids are provided to the fluid compartment in predefined mixing ratio by applying a pressure difference between the first and second fluid ports and at least one exit port; applying a pulsed electric field between the first and second electrodes thereby electroporating the cells with at least one compound; and removing the fluids from the fluid compartment; and applying a ground potential to the cell suspension by the third and the fourth electrodes disposed at the first fluid inlet port and the at least one exit port.

10. The process according to claim 9 , wherein the first and second fluids are provided to the fluid compartment to a predefined filling level; and wherein the filling level of the fluid compartment is determined by measuring the capacitance between the first and second electrode.

11. The process according to claim 9 wherein after removing the fluids from the fluid compartment, the electroporated cells are detected and depleted from the fluids.

12. The process according to claim 9 , wherein after removing the fluids from the fluid compartment, the electroporated cells are cultivated.

13. A system for electroporation of cells comprising the disposable cartridge according to claim 1 , a first storage container containing a cell suspension; a second storage container containing a fluid comprising at least one compound to be electroporated into the cells; a third storage container for electroporated cells; a pump and/or a vacuum source; and a tubing set providing fluid communication between the first, second and third storage containers and the disposable cartridge, wherein the system is closed to outside atmosphere.

14. The system according claim 13 , wherein the third storage container is configured as a cultivation chamber.

15. The disposable cartridge of claim 1 , wherein the voltage pulse is at least one of exponentially decaying, square wave, and combinations thereof.

16. A method for creating pores in a cell membrane, comprising:
providing the disposable cartridge of claim 1 ,
filling the disposable cartridge of claim 1 with a suspension of cells;
mixing the suspension; and
electroporating the cells by applying a pulse of at least 2 kV/cm to create pores in the cell membranes.

17. The disposable cartridge of claim 1 , wherein the first and second electrodes fit conformally within the fluid compartment, wherein both the fluid compartment and the first and second electrodes have acute and obtuse angles.

18. The disposable cartridge of claim 1 , wherein both the first and second electrodes and the fluid compartments have a shape of a trapezoid or parallelogram, with the trapezoid of the first and second electrodes fitting within the trapezoid of the fluid compartment.

Assignees

• Miltenyi Biotec Gmbh

Inventors

• Miltenyi Stefan
• Boddenberg Jan
• Kabaha Eiad
• Peters Ralf-Peter

Classifications

• C12N15/87

  Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation · CPC title

• C12M35/02 Primary

  Electrical or electromagnetic means, e.g. for electroporation or for cell fusion · CPC title

• C12M23/28

  disposable or single use · CPC title

• C12M23/16 Primary

  Microfluidic devices; Capillary tubes (integrated microfluidic structures B01L3/5027; microreactors B01J19/0093) · CPC title

• C12N13/00

  Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves · CPC title