Methods and apparatus for the manipulation of particle suspensions and testing thereof

• US 8,293,524 B2
• Filed: 03/25/2007
• Issued: 10/23/2012
• Est. Priority Date: 03/31/2006
• Status: Expired due to Fees

• Current Assignee: Fluxion Biosciences Incorporated
• Original Assignee: Fluxion Biosciences Incorporated
• Inventors: Ionescu-Zanetti, Cristian, Khine, Michelle, Schwartz, Michael, Blatz, Andrew
• Primary Examiner(s): Marcheschi, Michael
• Assistant Examiner(s): Doe, Shanta G
• Application Number:
  US11/690,831
• Publication Number:
  US 20070243523A1
• Time in Patent Office: 2,039 Days
• Field of Search: 435/287.1, 435/288.5, 435/305.2, 435/286.2, 435/287.3, 435/297.1, 435/297.2, 435/287.2, 435/395, 422/82.01
• US Class Current: 435/297.2
• CPC Class Codes: B01L 2200/025 Align devices or objects to ensure defined positions relative to each other
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2200/0668 Trapping microscopic beads
• CPC Class Codes: B01L 2200/0689 Sealing
• CPC Class Codes: B01L 2300/041 Connecting closures to device or container
• CPC Class Codes: B01L 2300/046 Function or devices integrated in the closure
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0829 Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0867 Multiple inlets and one sample wells, e.g. mixing, dilution
• CPC Class Codes: B01L 2300/163 Biocompatibility
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/5025 for parallel transport of multiple samples
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/50273 characterised by the means or forces applied to move the fluids micropumps F04B19/006, of the membrane type F04B43/043
• CPC Class Codes: B01L 3/502738 : characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: B01L 3/502761 : specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: B01L 3/565 : Seals in general F16L
• CPC Class Codes: C12M 21/06 : for in vitro fertilization
• CPC Class Codes: C12M 23/16 : Microfluidic devices; Capillary tubes integrated microfluidic structures B01L3/5027; microreactors B01J19/0093
• CPC Class Codes: C12M 35/04 : Mechanical means, e.g. sonic waves, stretching forces, pressure or shear stimuli
• CPC Class Codes: C12M 41/36 : of biomass, e.g. colony counters or by turbidity measurements electrooptical investigation of individual particles G01N15/14, flow cytometers G01N15/1404
• CPC Class Codes: G01N 15/1404 : Handling flow, e.g. hydrodynamic focusing
• CPC Class Codes: G01N 15/1409 : Handling samples, e.g. injecting samples
• CPC Class Codes: G01N 15/1433 : using image recognition
• CPC Class Codes: G01N 15/1459 : the analysis being performed on a sample stream
• CPC Class Codes: G01N 15/1484 : microstructural devices
• CPC Class Codes: G01N 2015/1006 : for cytology
• CPC Class Codes: G01N 2015/1027 : Determining speed or velocity of a particle
• CPC Class Codes: G01N 2015/1495 : Deformation of particles
• CPC Class Codes: G01N 2015/1497 : Particle shape
• CPC Class Codes: G01N 33/48728 : Investigating individual cells, e.g. by patch clamp, voltage clamp investigating individual particles in general G01N15/10

Abstract

Apparatus and methods are provided for analysis of individual particles in a microfluidic device. The methods involve the immobilization of an array of particles in suspension and the application of experimental compounds. Such methods can also include electrophysiology studies including patch clamp recording, electroporation, or both in the same microfluidic device. The apparatus provided includes a microfluidic device coupled to a multi-well structure and an interface for controlling the flow of media within the microchannel device.

Claims

1. A system comprising:

a microfluidic device having a plurality of trapping areas configured to trap a cell, wherein each trapping area includes a first channel and a plurality of second channels intersecting the first channel, wherein the first channel and the plurality of second channels are horizontal channels defined in part by a common surface, and wherein the first channel has a first height and the plurality of second channels have a second height, less than the first height, wherein at least one of the plurality of second channels is a horizontal channel configured to connect the first channel to a reservoir;

a well plate bonded to the microfluidic device, wherein the well plate defines a plurality of wells, wherein a respective grouping of the plurality of wells is associated with each of the trapping areas, and wherein each respective grouping includes a first well in fluid communication with the first channel and a plurality of second wells each in communication with a respective one of the plurality of second channels; and

an interface coupled to the well plate, wherein the interface is configured for individually addressing the first well and the plurality of second wells within at least one of the respective groupings, and wherein the interface is configured to apply pneumatic pressures to the first well and at least one of the second wells sufficient to move a fluid within the microfluidic device.

2. The system of claim 1, wherein the well plate comprises a microplate, the microfluidic device is integrated into the microplate and the microplate is selected from the group consisting of a 24-well, 96-well, 384-well, and a 1536-well microplate.

3. The system of claim 1, wherein the interface is coupled to at least a section of the well plate.

4. The system of claim 1, wherein the trapping areas comprise regions of the microfluidic device adapted for immobilizing a plurality of cells.

5. The system of claim 1, wherein the trapping areas comprise a region of the microfluidic device that is optically accessible.

6. The system of claim 1, wherein the trapping areas comprise a region of the microfluidic device that is optically accessible microscopically during patch clamp measurements.

7. The system of claim 1, further comprising an electrode array in electronic communication with the microfluidic device, wherein the electrode array comprises electrodes, and wherein at least one electrode extends into at least one of said wells when the well plate and interface are coupled.

8. The system of claim 7, wherein the electrodes are substantially cylindrical.

9. The system of claim 8, wherein the substantially cylindrical electrodes comprise an end, wherein a section cut out of the end of at least one electrode extends at least partially into the microfluidic device.

10. The system of claim 7, wherein the electrode array is adapted for electrophysiological analysis of plurality of cells.

11. The system of claim 10, wherein the electrophysiological analysis comprises recording selected from the group consisting of whole-cell recording and patch clamp recording.

12. The system of claim 7, wherein the interface is adapted to act as a shield for ambient electromagnetic waves.

13. The system of claim 7, wherein the interface is connectable to a patch clamp amplifier.

14. The system of claim 1, wherein the interface provides an aperture for optical access.

15. The system of claim 1, further comprising a current measurement system in electrical communication with the microfluidic device.

16. The system of claim 1, wherein said plurality of second channels intersect said first channel at respective lateral openings for immobilizing particles, wherein the distance between said lateral openings is substantially below 0.1 mm.

17. The system of claim 16, wherein the particles are selected from cells, vesicles and oocytes.

18. The system of claim 17, wherein the particles are cells.

19. The system of claim 18, wherein a plurality of the cells can be simultaneously observed microscopically.

20. The system of claim 19, further comprising one or more compound streams in fluid communication with the immobilized cells wherein the plurality of the cells is exposed to the same compound stream.

21. The system of claim 1, wherein said well plate comprises at least a portion of a wellplate having at least 96 wells, and wherein said interface is configured to provide at least 96 simultaneous pressure connections to individual ones of said 96 wells.

22. The system of claim 1, wherein said interface comprises a detachable interface configured to reversibly mate to said well plate.

23. The system of claim 1, wherein said interface comprises a gasket configured to at least partially cover said well plate and tubing configured to transport a gas into the at least one reservoir.

24. A system comprising:

a microfluidic device comprising;

a substrate;

a main flow channel having a first height in said substrate adapted to hold cells in a fluidic medium;

at least one lateral opening in a side of said main flow channel;

at least one horizontal trapping channel in said substrate operatively connected to said lateral opening, wherein the horizontal trapping channel has a second height less than the first height, and wherein the main flow channel, the at least one lateral opening, and the at least one horizontal trapping channel are defined in part by a same surface; and

at least two electrical connections on a same side of said substrate, one connected to said main flow channel and one connected to said trapping channel;

wherein a particle in the main flow channel can be immobilized at said lateral opening by negative pressure in the trapping channel;

at least one reservoir in fluid communication with the microfluidic device, wherein said at least one reservoir comprises at least a portion of a wellplate, wherein the wellplate is bonded to the microfluidic device; and

a pneumatic interface coupled to the microfluidic device, wherein the interface is adapted for moving material within the microfluidic device at least in part by applying a pressure to an air-fluid interface in the reservoir, wherein said interface is configured to provide simultaneous pressure connections to individual wells of the wellplate.

25. The system of claim 24, wherein the particles are selected from the group consisting of cells, vesicles and oocytes.

26. The system of claim 25, wherein the particles are cells.

27. The system of claim 26, wherein a plurality of the cells can be simultaneously observed microscopically.

28. The system of claim 25, further comprising one or more compound streams in fluid communication with the immobilized cells wherein a plurality of the cells is exposed to the same compound stream.

29. The system of claim 26, wherein the detected characteristic of a cell is an ion-flux activity.

30. The system of claim 29, wherein the ion-flux activity is hERG activity.

31. The system of claim 24, wherein the main flow channel comprises an array of lateral openings, and the distance between lateral openings is substantially below 0.1 mm.

32. The system of claim 24, wherein said at least a portion of a wellplate has at least 96 wells, and wherein said interface is configured to provide at least 96 simultaneous pressure connections to individual ones of said 96 wells.

33. The system of claim 24, wherein said interface comprises a detachable interface configured to reversibly mate to said at least one reservoir.

34. The system of claim 24, wherein said interface comprises a gasket configured to at least partially cover said at least one reservoir and tubing configured to transport a gas into the at least one reservoir.