Microwell arrays with nanoholes

• US 20060240543A1
• Filed: 04/05/2004
• Published: 10/26/2006
• Est. Priority Date: 04/03/2003
• Status: Active Grant

• Current Assignee: University of Washington
• Original Assignee: University of Washington
• Inventors: Kosar, Turgut Fettah, Folch, Albert
• Granted Patent: US 7,501,279 B2
• Time in Patent Office: Days
• US Class Current: 435/288.300
• CPC Class Codes: B01L 2200/0668 Trapping microscopic beads
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 2400/0409 centrifugal forces
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/043 magnetic forces
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 3/50255 Multi-well filtration
• 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 23/16 Microfluidic devices; Capillary tubes integrated microfluidic structures B01L3/5027; microreactors B01J19/0093
• CPC Class Codes: C12M 25/04 in combination with well or multiwell plates, i.e. culture inserts
• CPC Class Codes: G01N 33/48728 Investigating individual cells, e.g. by patch clamp, voltage clamp investigating individual particles in general G01N15/10

Abstract

A device for conducting parallel analysis or manipulation of multiple cells or biomolecules is disclosed. In one embodiment, the device comprises a silicon chip with a microwell, and at least one membrane suspended at the bottom opening of the microwell. The suspended portion of the membrane defines a nanohole that provides access to the material on the other side of the membrane.

Claims

1. A device comprising:

a substrate defining at least one microwell adapted to receive fluid, wherein the microwell comprises a bottom opening,

a membrane suspended across the microwell bottom opening and wherein the suspended portion defines a nanohole, and

a channel layer comprising at least one microchannel, positioned such that the microchannel is in fluid communication with a fluid in the microwell via the nanohole.

2. The device as claimed in claim 1, wherein the nanohole is sized to permit cells or biomolecules smaller than a designated size through the membrane while retaining larger cells or biomolecules behind the membrane.

3. The device as claimed in claim 1, wherein the nanohole has a diameter ranging in size from about 1 nanometer to about 50 microns.

4. The device as claimed in claim 1, wherein the membrane comprises silicon nitride.

5. The device as claimed in claim 1, wherein the membrane comprises silicon dioxide.

6. The device as claimed in claim 1, wherein the channel layer comprises a polymer.

7. The device as claimed in claim 1, wherein the channel layer comprises poly(dimethylsiloxane).

8. The device as claimed in claim 1, wherein the nanohole is sized to permit a single strand of DNA or protein to pass through the nanohole.

9. The device as claimed in claim 1, wherein the nanohole measures between about 1 to about 3 nanometers in diameter.

10. A method for providing access to a cell or biomolecule comprising:

obtaining a device for containing a cell or biomolecule, wherein the device comprises a microwell with a bottom opening, a membrane with a portion suspended across the bottom opening, wherein the suspended portion of the membrane defines a nanohole, and

positioning the cell or biomolecule on one side of the suspended portion of the membrane, such that the cell or biomolecule can be accessed via the nanohole.

11. The method of claim 10, wherein the device further comprises a microchannel layer, wherein the microchannel layer defines at least one microchannel with an opening under the nanohole.

12. The method of claim 10, wherein the microchannel provides access to the cell or biomolecule.

13. The method of claim 10, further comprising applying suction to the side of the suspended portion of the membrane opposite the cell or biomolecule to position the cell or biomolecule at the nanohole.

14. The method of claim 10, wherein the device is adapted to enable the cell or biomolecule to be automatically guided to the nanohole when suction is applied to the other side of the nanohole to position cell or biomolecule.

15. The method of claim 10, wherein the positioning step is achieved via the microwell.

16. The method of claim 10, wherein the positioning step is achieved via the microchannel.

17. A device comprising:

a substrate defining at least one microwell, wherein the microwell comprises a bottom opening,

a membrane layer defining a nanohole, wherein the nanohole is positioned below the bottom opening of the microwell,

a polymeric insulator positioned below the membrane layer, wherein the insulator defines a channel positioned below the nanohole in the membrane layer,

wherein the membrane layer, the microchannel and the microwell bottom opening are configured such that a fluid contained in the microchannel contacts the membrane layer only at a portion of the membrane which is below the microwell bottom opening.

18. The device as claimed in claim 17, wherein the configuration of the membrane layer, the microchannel and the microwell bottom opening prevents fluid contained in the microchannel from contacting a portion of the membrane layer which is below the substrate.

19. The device as claimed in claim 17, wherein the microchannel has a portion with an area below the microwell bottom opening, which is smaller than the area defined by the perimeter of the microwell bottom opening.

20. A method of manufacturing a device for providing access to cells or biomolecules comprising:

obtaining a substrate with a top side and a bottom side,

forming a membrane layer on the substrate,

forming a window in the membrane layer on the top side of the substrate and etching a microwell in the substrate at the window,

forming a nanohole in the membrane layer, wherein the nanohole is positioned on the bottom side of the substrate under the microwell,

providing at least one channel in a channel layer attached to the bottom side of the substrate with the channel having an opening positioned under the nanohole.

21. A method of manufacturing a device for providing access to cells or biomolecules comprising:

obtaining a silicon substrate with a top side and a bottom side,

forming a silicon nitride layer on the substrate,

depositing a metal layer above the silicon nitride layer and creating photoresist islands using a photoresist polymer,

etching at least one nanohole in the silicon nitride layer by reactive ion etching, using a film containing a nanohole as an etch mask,

lithographically etching at least one microwell window in the silicon nitride layer, wherein the position of the microwell window is positioned opposite the nanohole, using the photoresist islands as a guide,

anisotropically etching a microwell at the microwell window,

obtaining a channel layer comprising at least one channel,

sealing the channel layer to the silicon nitride such that the channel is positioned beneath the nanohole.