Method and apparatus for high throughput high efficiency transfection of cells

Patent metadata

Field	Value
Publication number	US-11859162-B2
Application number	US-201916557820-A
Country	US
Kind code	B2
Filing date	Aug 30, 2019
Priority date	Aug 31, 2018
Publication date	Jan 2, 2024
Grant date	Jan 2, 2024

Abstract

Transfer of genetic and other materials to cells is conducted in a hands-free, automated, high throughput, continuous process. A system using a microfluidic hydrodynamic sheath flow configuration includes arrangements for pushing cells from side streams containing a cell culture medium to a central stream containing an electroporation buffer. Electroporation can be conducted in an assembly in which two or more microfluidic channels are provided in a parallel configuration and in which various layers can be stacked together to form a laminate type structure.

First claim

What is claimed is:

1. A method for introducing a payload into cells, the method comprising:
directing cells from a first incubator to an assembly that contains multiple microfluidic devices;
acoustically driving cells from a cell culture medium flowing in a side stream of a sheath flow configuration, to an electroporation medium flowing through a central stream of the sheath flow configuration;
applying an electric field to cells in the electroporation medium; and
transferring or allowing the transfer of the payload into the cells.

2. The method of claim 1 , further comprising driving cells containing the payload from the electroporation fluid into a cell culture fluid.

3. The method of claim 2 , further comprising storing cells containing the payload in a second incubator.

4. The method of claim 1 , further comprising administering cells containing the payload to a subject in need of diagnosis, prophylaxis or treatment.

5. The method of claim 1 , wherein the payload is supplied in the electroporation medium.

6. The method of claim 1 , further comprising driving cells containing the payload from the electroporation medium to a third medium.

7. The method of claim 1 , wherein the electric field is applied by a pair of electrodes provided for each microfluidic device in the assembly.

8. A method for manufacturing cells for immunotherapy, the method comprising:
acoustically transferring cells from a first buffer, wherein the first buffer is a cell culture medium, into an electroporation buffer;
permeabilizing the cells by electroporation;
allowing a payload to transfer into the permeabilized cells; and
transferring the cells containing the payload into a second buffer, wherein, the method is conducted in an automated and continuous flow mode, wherein the electroporation buffer forms a central stream and the first buffer forms a side stream of a sheath flow configuration, and wherein, the throughput is at least 4 million cells per minute.

9. The method of claim 8 , wherein the method is conducted in an assembly comprising multiple microfluidic devices.

10. A method for introducing a payload into cells, the method comprising:
flowing an electroporation medium as a central stream of a sheath flow configuration;
flowing a first medium containing cells as a side stream in the sheath flow configuration;
applying acoustic energy to drive cells from the first medium to the electroporation medium;
applying an electric field to permeabilize the cells in the electroporation medium; and
allowing the payload to transfer into the permeabilized cells.

11. The method of claim 10 , further comprising driving the cells containing the payload from the electroporation medium to the first medium.

12. The method of claim 10 , further comprising driving the cells containing the payload from the electroporation medium to a second medium, wherein the second medium is a central stream in a sheath flow configuration.

13. The method of claim 10 , further comprising applying a second acoustic energy to drive cells containing the payload from the electroporation medium to a second medium flowing as a center stream in a sheath flow configuration.

14. The method of claim 10 , wherein the method is conducted in a microfluidic device or in an assembly comprising multiple microfluidic devices.

15. The method of claim 10 , wherein the first medium containing cells is supplied from an incubator.

16. The method of claim 10 , wherein the payload is provided in the electroporation medium.

17. The method of claim 10 , wherein the method is conducted in an automated continuous flow mode.

18. The method of claim 10 , further comprising administering the cells containing the payload to a patient in need of diagnosis, prophylaxis or treatment.

Assignees

• Charles Stark Draper Laboratory Inc

Inventors

• Tandon Vishal
• Lissandrello Charles A
• Balestrini Jenna L
• Coppeta Jonathan R
• Swierk Patricia A

Classifications

• C12M23/16 Primary

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

• C12M35/02

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

• B01L3/50273

  characterised by the means or forces applied to move the fluids · CPC title

• B01L3/502715

  characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces · CPC title

• B01L2300/047

  Additional chamber, reservoir · CPC title