Bioreactors with multiple chambers

• US 7,977,089 B2
• Filed: 08/27/2003
• Issued: 07/12/2011
• Est. Priority Date: 08/27/2002
• Status: Expired due to Fees

• Current Assignee: Vanderbilt University
• Original Assignee: Vanderbilt University
• Inventors: Leboeuf, Eugene J., Prokop, Ales, Cliffel, David, Wikswo, John P., Reiserer, Randall S., Haselton, Frederick R., Stremler, Mark A., Baudenbacher, Franz J.
• Primary Examiner(s): BOWERS, NATHAN ANDREW
• Application Number:
  US10/525,549
• Publication Number:
  US 20060166354A1
• Time in Patent Office: 2,876 Days
• Field of Search: 435/288.7, 435/808, 435/288.4, 435/288.5, 435/297.1, 435/297.5, 435/305.1, 435/305.2, 205/777.5, 205/778, 205/779, 204/403.01
• US Class Current: 435/305.2
• CPC Class Codes: B01L 2200/10 Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0681 Filter
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/163 Biocompatibility
• CPC Class Codes: B01L 2400/0472 Diffusion
• CPC Class Codes: B01L 3/5027 by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: C12M 23/12 Well or multiwell plates C12M25/04 takes precedence
• CPC Class Codes: C12M 23/16 Microfluidic devices; Capillary tubes integrated microfluidic structures B01L3/5027; microreactors B01J19/0093
• CPC Class Codes: C12M 29/10 Perfusion
• CPC Class Codes: C12M 41/00 Means for regulation, monitoring, measurement or control, e.g. flow regulation controlling or regulating chemical, physical or physicochemical processes B01J19/0006; heating or cooling apparatus for laboratory use B01L7/00; electro optical investigation of individual particles, flow cytometers G01N15/14; automatic analysis G01N35/00; controlling or regulating in general G06N
• CPC Class Codes: Y10S 435/808 Optical sensing apparatus

Abstract

A bioreactor for cultivating living cells in a liquid medium. In one embodiment of the present invention, the bioreactor has a first substrate having a first surface and an opposite second surface, defining a chamber therebetween for receiving the cells and the liquid medium. The bioreactor further has a barrier dividing the chamber into a first subchamber and a second subchamber, wherein the barrier has a porosity to allow the first subchamber and the second subchamber in fluid communication and allow at least one predetermined type of cells to permeate between the first subchamber and the second subchamber.

Claims

1. A bioreactor for cultivating living cells in a liquid medium comprising:

(a) a first substrate having a first surface and an opposite second surface, defining a chamber therebetween for receiving and culturing the cells and receiving the liquid medium;

(b) a barrier dividing the chamber into a first subchamber and a second subchamber, wherein the barrier comprises a porous material and has a porosity to allow the first subchamber and the second subchamber to be in fluid communication and allow at least one predetermined type of cells to permeate between the first subchamber and the second subchamber;

(c) a second substrate positioned adjacent to the first surface of the first substrate;

(d) a third substrate, wherein the third substrate is positioned adjacent to the second surface of the first substrate;

(e) an electrochemical measuring system positioned in the third substrate and adapted for electrochemical measurements of the cells responsive to the liquid medium in at least one of the first subchamber and the second subchamber;

(f) a fourth substrate positioned above the third substrate such that the third substrate forms an intermediate layer between the first substrate and the fourth substrate; and

(g) means positioned in the fourth substrate and adapted for optical measurements of the cells responsive to the liquid medium in at least one of the first subchamber and the second subchamber.

2. The bioreactor of claim 1, wherein the first subchamber is adapted for receiving a first type of material and the second subchamber is adapted for receiving a second type of material.

3. The bioreactor of claim 2, wherein each of the first type of material and the second type of material contains at least one selected from the group of cells, chemicals, and fluids.

4. The bioreactor of claim 3, wherein the cells comprise bacteria.

5. The bioreactor of claim 3, wherein the cells comprise protozoa.

6. The bioreactor of claim 1, further comprising a biocompatible coating layer applied to the chamber walls.

7. The bioreactor of claim 6, wherein the biocompatible coating layer comprises a material that may inhibit cell adhesion to the biocompatible coating layer, enhance cell adhesion to the biocompatible coating layer, or function as a fluorescent marker or indicator of the state of cells.

8. The bioreactor of claim 1, further comprising at least one inlet port and an input transfer channel in fluid communication with the inlet port and one of the first subchamber and the second subchamber for allowing delivery of the cells, fluids or chemicals to the corresponding subchamber.

9. The bioreactor of claim 8, further comprising at least one outlet port and an outlet transfer channel in fluid communication with the outlet port and one of the first subchamber and the second subchamber for allowing removal of the cells, fluids or chemicals from the corresponding subchamber.

10. The bioreactor of claim 9, further comprising at least one auxiliary port and an auxiliary channel in fluid communication with the auxiliary port and one of the input transfer channel and the outlet transfer channel for flushing the corresponding transfer channel.

11. The bioreactor of claim 10, further comprising at least one access port and an access channel in fluid communication with the access port and one of the first subchamber and the second subchamber for allowing delivery or removal of the cells, fluids, chemicals, coating material or sensing material to the corresponding subchamber.

12. The bioreactor of claim 11, wherein the second substrate defines a plurality of connection channels, each of the connection channels being formed so as to be in fluid communication with a corresponding one of the inlet port, the outlet port, the auxiliary port, and the access port.

13. The bioreactor of claim 12, further comprising a plurality of connection ports, each of the connection ports being formed with a channel and being positioned to the second substrate such that each channel of the connection ports is in fluid communication with a corresponding one of the connection channels formed in the second substrate.

14. The bioreactor of claim 1, wherein the first substrate is fabricated from glass, Mylar, PDMS, silicon, a polymer, a semiconductor, or any combination of them.

15. The bioreactor of claim 1, wherein the barrier is microfabricated so as to form a structure allowing the fluid communication between the first subchamber and the second subchamber.

16. The bioreactor of claim 1, wherein the means for electrochemical measurements comprises:

(i) a reference electrode;

(ii) a counter electrode;

(iii) a plurality of edge connector pads; and

(iv) a plurality of electrically conductive leads, where a first electrically conductive lead electrically couples the reference electrode to a corresponding edge connector pad, and a second electrically conductive lead electrically couples the counter electrode to a corresponding edge connector pad.

17. The bioreactor of claim 16, wherein the means for electrochemical measurements further comprises:

a plurality of individually addressable working electrodes, each being electrically coupled to a corresponding edge connector pad through a corresponding electrically conductive lead.

18. The bioreactor of claim 17, wherein the liquid medium comprises at least one analyte, and wherein the plurality of individually addressable working electrodes are adapted for sensing the concentration of a single analyte of the liquid medium at multiple locations in the chamber or the concentrations of a plurality of analytes of the liquid medium at multiple locations in the chamber at a time period shorter than a characterization reaction time related to at least one of cellular physiological activities of the cells.

19. The bioreactor of claim 18, wherein the plurality of individually addressable working electrodes are further adapted for measuring the metabolic variables related to the cells responsive to the liquid medium at multiple locations in the chamber at a time period shorter than a characterization reaction time related to at least one of cellular physiological activities of the cells.

20. The bioreactor of claim 1, wherein the third substrate comprises a semiconductor material.

21. The bioreactor of claim 20, wherein the semiconductor material comprises silicon.

22. The bioreactor of claim 1, wherein the means for optical measurements comprises:

(i) a plurality of optical sensors;

(ii) a plurality of edge connector pads; and

(iii) a plurality of optically conductive leads, each optically coupling an optical sensor to a corresponding edge connector pad.

23. The bioreactor of claim 22, wherein the plurality of optical sensors comprises at least one device selected from the group of an LED and photodiode pair, a fiber optic coupler, and an optical detecting head.

24. The bioreactor of claim 23, wherein the liquid medium comprises at least one analyte, and wherein the plurality of optical sensors are adapted for sensing the concentration of a single analyte of the liquid medium at multiple locations in the chamber or the concentrations of a plurality of analytes of the liquid medium at multiple locations in the chamber at a time period shorter than a characterization reaction time related to at least one of cellular physiological activities of the cells.

25. The bioreactor of claim 24, wherein the plurality of optical sensors are further adapted for measuring the metabolic variables related to the cells responsive to the liquid medium at multiple locations in the chamber at a time period shorter than the characterization reaction time related to at least one of cellular physiological activities of the cells.

26. The bioreactor of claim 1, wherein the fourth substrate comprises a semiconductor material.

27. The bioreactor of claim 26, wherein the fourth substrate is at least partially transparent.