Microfabricated device and method for multiplexed electrokinetic focusing of fluid streams and a transport cytometry method using same

• US 6,120,666 A
• Filed: 06/16/1998
• Issued: 09/19/2000
• Est. Priority Date: 09/26/1996
• Status: Expired due to Term

• Current Assignee: Lockheed Martin Energy Research Corporation (Martin Marietta Corporation)
• Original Assignee: UT-Battelle LLC
• Inventors: Ramsey, J. Michael, Jacobson, Stephen C.
• Primary Examiner(s): Warden, Jill
• Assistant Examiner(s): Starsiak, John S.
• Application Number:
  US09/098,178
• Time in Patent Office: 826 Days
• Field of Search: 204/450, 204/451, 204/452, 204/453, 204/454, 204/455, 204/600, 204/601, 204/602, 204/603, 204/604, 204/605, 422/100, 422/68.1, 422/81, 137/597, 137/594, 137/816, 137/818, 137/825, 137/806, 137/807, 73/863, 73/864.15, 222/564, 435/287.1, 435/287.2, 435/287.3, 435/288.4, 435/288.5, 435/288.6, 436/180
• US Class Current: 204/452
• CPC Class Codes: B01F 33/30 Micromixers
• CPC Class Codes: B01F 33/3011 using a sheathing stream of a fluid surrounding a central stream of a different fluid, e.g. for reducing the cross-section of the central stream or to produce droplets from the central stream
• CPC Class Codes: B01F 33/3031 using electro-hydrodynamic [EHD] or electro-kinetic [EKI] phenomena to mix or move the fluids
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/0418 electro-osmotic flow [EOF]
• CPC Class Codes: B01L 2400/0421 electrophoretic flow
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 3/502738 characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: B01L 3/502776 specially adapted for focusing or laminating flows
• CPC Class Codes: G01N 15/1404 Handling flow, e.g. hydrodynamic focusing
• 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 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 27/44721 by optical means
• CPC Class Codes: G01N 27/44773 Multi-stage electrophoresis, e.g. two-dimensional electrophoresis
• CPC Class Codes: G01N 27/44791 Microapparatus sample containers with integrated microfluidic structures B01L3/5027
• CPC Class Codes: G01N 30/6095 : Micromachined or nanomachined, e.g. micro- or nanosize
• CPC Class Codes: Y10S 366/01 : Micromixers: continuous laminar flow with laminar boundary mixing in the linear direction parallel to the fluid propagation with or without conduit geometry influences from the pathway
• CPC Class Codes: Y10T 137/2082 : Utilizing particular fluid

Abstract

A microfabricated device and method for electrokinetic transport of a liquid phase biological or chemical material is described. In accordance with one aspect of the present invention there is provided a microchip that is adapted for the simultaneous spatial confinement of electrokinetically driven fluidic material streams on a substrate. The apparatus includes a focusing chamber formed in a surface of the substrate and in fluid communication with two sample fluid channels and three focusing fluid channels. The device further includes electromotive means operatively connected to the sources of the sample fluid and the source of focusing fluid for electrokinetically driving the respective streams of the sample and focusing fluids through the respective channels into the focusing chamber such that the focusing fluid streams spatially confine the first and second sample fluid streams within the focusing chamber. In accordance with another aspect of this invention, there is provided a cytometry method for analyzing microscopic particles in a fluid medium on a microchip by utilizing the focusing function of the microchip. In the disclosed cytometry process the width of the fluid stream is narrowed in the focusing chamber. The microscopic particles in the focused sample fluid are then detected and/or measured using light scattering or other techniques.

Claims

1. An apparatus for the simultaneous spatial confinement of material streams on a substrate, comprising:

a focusing chamber formed in a surface of the substrate;

first and second sample channels formed in the surface of said substrate for carrying sample fluid streams therethrough, said sample channels having respective first ends in fluid communication with a source of a sample fluid and respective second ends in fluid communication with said focusing chamber;

first, second, and third focusing channels formed in the surface of said substrate for carrying focusing fluid streams therethrough, said focusing channels being formed such that said first sample channel is positioned between said first and second focusing channels and said second sample channel is positioned between said second and third focusing channels, said focusing channels having respective first ends in fluid communication with a source of focusing fluid and respective second ends in fluid communication with said focusing chamber;

first and second waste channels formed in the surface of said substrate, said waste channels having respective first ends in fluid communication with said focusing chamber and having respective second ends in fluid communication with a waste reservoir; and

means operatively connected to the source of the sample fluids and the source of focusing fluid for driving the respective streams of the sample and focusing fluids through the respective channels into said focusing chamber such that the focusing fluid streams spatially confine the first and second sample fluid streams within said focusing chamber.

2. An apparatus according to claim 1 further comprising:

first and second sample collection channels formed in the substrate for carrying the respective streams of the sample fluid, said sample collection channels having respective first ends in fluid communication with corresponding waste channels and having respective second ends in fluid communication with first and second collection reservoirs, respectively; and

a buffer channel formed in the substrate for carrying a stream of buffer fluid, said buffer channel having a first end in fluid communication with a source of buffer fluid and a second end in fluid communication with said first and second waste channels; and

means operatively connected to the source of the buffer fluid, the collection reservoir, and the waste reservoir for driving the buffer fluid through the buffer channel to direct the streams of the sample fluid to corresponding waste channels, to corresponding collection channels, or to a combination of a waste channel and a collection channel.

3. An apparatus as set forth in claim 1 wherein the respective first ends of the first and second waste channels are positioned substantially opposite the respective second ends of the first and second sample channels in said focusing chamber.

4. An apparatus as set forth in claim 1 wherein the first and second waste channels have respective widths that are narrower in dimension than the widths of the first and second sample channels, respectively.

5. An apparatus as set forth in claim 1 wherein the first and second sample channels have respective widths that are narrower in dimension than the widths of the first and second waste channels, respectively.

6. An apparatus as set forth in claim 1 wherein the widths of the sample channels and the widths of the waste channels are narrower in dimension than the widths of the focusing channels.

7. A method for analyzing microscopic particles in a fluid medium on a microchip comprising the steps of:

conducting a stream of a sample fluid containing microscopic particles through a sample fluid channel to a focusing chamber;

narrowing the width of the sample fluid stream in the focusing chamber by transporting a focusing fluid into the focusing chamber such that the width of the sample fluid stream in the focusing chamber corresponds substantially to a detection zone; and

detecting the microscopic particles in the focused sample fluid in the detection zone.

8. A method as set forth in claim 7, further comprising the step of measuring a physical characteristic of the microscopic particles.

9. A method as set forth in claim 8 further comprising the steps of:

comparing the measured physical characteristic to a reference value for the physical characteristic;

diverting the sample fluid stream to a first channel when the measured physical characteristic has a first quantitative relationship to the reference value; and

diverting the sample fluid stream to a second channel when the measured physical characteristic has a second quantitative relationship to the reference value.

10. A method as set forth in claim 7 wherein the step of narrowing the width of the sample fluid stream further comprises the step of narrowing the width of the sample fluid channel.

11. Apparatus for analyzing microscopic particles in a fluid medium on a microchip having a substrate, comprising:

a focusing chamber formed in a surface of the substrate;

a sample channel formed in the surface of said substrate for carrying a sample fluid stream therethrough, said sample channel having a first end in fluid communication with a source of a sample fluid containing microscopic particles and a second end in fluid communication with said focusing chamber;

first and second focusing channels formed in the surface of said substrate for carrying focusing fluid streams therethrough, said focusing channels having respective first ends in fluid communication with a source of focusing fluid and respective second ends in fluid communication with said focusing chamber;

a detection channel formed in the surface of said substrate and having a first end in fluid communication with said focusing chamber for receiving the sample fluid therefrom;

means operatively connected to the source of the sample fluid and the source of focusing fluid for driving the respective streams of the sample and focusing fluids through the respective channels into said focusing chamber such that the focusing fluid streams spatially confine the sample fluid stream within said focusing chamber and said detection channel; and

means for detecting the microscopic particles in the spatially confined sample fluid in said detection channel.

12. An apparatus as set forth in claim 11 further comprising a collection channel formed in the surface of said substrate, said collection channel having a first end in fluid communication with said detection channel.

13. An apparatus as set forth in claim 12 further comprising a waste channel formed in the surface of said substrate, said waste channel having a first end in fluid communication with said detection channel.

14. An apparatus as set forth in claim 13 wherein the means for detecting the microscopic particles comprises analyzing means for measuring a physical characteristic of the microscopic particles.

15. An apparatus as set forth in claim 14 wherein the analyzing means comprises:

comparing means for comparing the measured physical characteristic to a reference value for the physical characteristic; and

diverting means for diverting the sample fluid stream to the collection channel when the measured physical characteristic has a first quantitative relationship to the reference value and for diverting the sample fluid stream to the waste channel when the measured physical characteristic has a second quantitative relationship to the reference value.

16. Apparatus as set forth in claim 13 further comprising

a buffer channel formed in the substrate for carrying a stream of buffer fluid, said buffer channel having a first end in fluid communication with a source of buffer fluid and a second end in fluid communication with said collection channel and said waste channel;

and

means operatively connected to the source of the buffer fluid, the collection channel and the waste channel for driving the buffer fluid through the buffer channel to direct the stream of the sample fluid to the waste channel, to the collection channel, or to both the waste channel and the collection channel.

17. An apparatus as set forth in claim 11 wherein the detection channel has a width that is narrower in dimension than the width of the sample channel.

18. An apparatus as set forth in claim 11 wherein the sample channel has a width that is narrower in dimension than the width of the detection channel.

19. An apparatus as set forth in claim 11 wherein the sample channel and the detection channel have respective widths that are narrower in dimension than the width of either of the focusing channels.

20. An apparatus for the spatial confinement of a material stream, comprising:

a focusing chamber formed in a surface of a substrate;

a first sample channel formed in the surface of the substrate for conducting a first sample fluid stream therethrough, said first sample channel having a first end in fluid communication with a source of a sample fluid and a second end in fluid communication with said focusing chamber;

first and second focusing channels formed in the surface of the substrate for conducting focusing fluid streams therethrough, said focusing channels having respective first ends in fluid communication with a source of focusing fluid and respective second ends in fluid communication with said focusing chamber;

a first waste channel formed in the surface of the substrate, said first waste channel having a first end in fluid communication with said focusing chamber and a second end in fluid communication with a waste reservoir; and

means operatively connected to the source of the sample fluid and the source of focusing fluid for driving the respective streams of the sample and focusing fluids through the focusing and sample channels into said focusing chamber, whereby the focusing fluid streams spatially confine the first sample fluid stream within said focusing chamber.

21. An apparatus as set forth in claim 20 wherein the first sample channel has a width that is narrower in dimension than the width of either of the focusing channels.

22. An apparatus as set forth in claim 20 wherein the first waste channel has a width that is narrower in dimension than the width of either of the focusing channels.

23. An apparatus as set forth in claim 22 wherein the first sample channel has a width that is narrower in dimension than the width of either of the focusing channels.