Assay device and method

• US 8,574,924 B2
• Filed: 04/28/2010
• Issued: 11/05/2013
• Est. Priority Date: 12/31/2003
• Status: Active Grant

• Current Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Sia, Samuel K., Linder, Vincent, Amir-Parviz, Babak, Siegel, Adam, Whitesides, George M.
• Primary Examiner(s): BROWN, MELANIE YU
• Application Number:
  US12/769,489
• Publication Number:
  US 20100279310A1
• Time in Patent Office: 1,287 Days
• Field of Search: 422/50, 422/55, 422/400, 422/68.1, 422/82.05, 435/7.1, 435/283.1, 435/287.1, 435/287.2, 436/518, 436/524, 436/525
• US Class Current: 436/525
• CPC Class Codes: B01L 2300/0822 Slides
• CPC Class Codes: B01L 2300/168 Specific optical properties, e.g. reflective coatings
• CPC Class Codes: B01L 3/502707 characterised by the manufacture of the container or its components by shaping or joining plastic parts B29C59/00, B29C65/00; by laminating B32B37/00; manufacture of microstructural devices in general B81C
• CPC Class Codes: G01N 2021/0346 Capillary cells; Microcells
• CPC Class Codes: G01N 21/05 Flow-through cuvettes G01N21/09 takes precedence; handling fluid samples G01N1/10
• CPC Class Codes: G01N 21/59 Transmissivity G01N21/25 takes precedence
• CPC Class Codes: G01N 33/5302 Apparatus specially adapted for immunological test procedures
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/56988 HIV or HTLV
• CPC Class Codes: G01N 33/585 with a particulate label, e.g. coloured latex
• CPC Class Codes: Y10T 436/11 Automated chemical analysis
• CPC Class Codes: Y10T 436/117497 with a continuously flowing sample or carrier stream
• CPC Class Codes: Y10T 436/118339 with formation of a segmented stream

Abstract

An assay method is described, which comprises the steps of immobilizing a binding partner (e.g., an antigen or antibody) for an analyte to be detected (e.g., an antibody or antigen) on a portion of a surface of a microfluidic chamber; passing a fluid sample over the surface and allowing the analyte to bind to the binding partner; allowing a metal colloid, e.g., a gold-conjugated antibody, to associate with the bound analyte; flowing a metal solution, e.g., a silver solution, over the surface such as to form an opaque metallic layer; and detecting the presence of said metallic layer, e.g., by visual inspection or by measuring light transmission through the layer, conductivity or resistance of the layer, or metal concentration in the metal solution after flowing the metal solution over the surface.

Claims

1. A method of determining a sample component comprising:

passing a fluid sample over a region of a microfluidic channel, the region comprising a maximum cross-sectional dimension of less than 1 mm;

allowing a sample component to bind with a binding partner disposed on a surface of the region;

allowing a metal colloid to associate with the sample component;

flowing a metal solution over the surface to form a metallic layer;

determining the formation of the metallic layer as a function of time; and

determining the sample component based on the formation of the metallic layer as a function of time.

2. The method of claim 1 wherein the metal colloid associates with a bound sample component.

3. The method of claim 1 wherein the metal colloid comprises gold.

4. The method of claim 2 wherein the metal colloid comprises a gold-conjugated antibody.

5. The method of claim 1 wherein the metallic layer comprises silver.

6. The method of claim 1 wherein the metal solution is a silver solution.

7. The method of claim 1 wherein the sample component is one of an antigen and an antibody and the binding partner is the other of the antigen and the antibody.

8. The method of claim 1 further comprising determining the opacity of the metallic layer.

9. The method of claim 8 wherein determining comprises irradiating the metallic layer with light and measuring light transmittance.

10. The method of claim 9 wherein the light is measured at the same wavelength at which it is transmitted.

11. The method of claim 1 further comprising measuring the conductivity of the metallic layer.

12. The method of claim 1 wherein the fluid is passed over a plurality of surfaces.

13. The method of claim 12 wherein each of the plurality of surfaces is associated with a different binding partner.

14. The method of claim 1 further comprising detecting the concentration of metal in the metal solution after flowing the metal solution over the surface.

15. The method of claim 1 wherein the sample comprises whole blood.

16. The method of claim 1, wherein the region of the microfluidic channel comprises at least one cross-sectional dimension of less than 100 microns.

17. The method of claim 1, wherein the binding partner is disposed across a width of the microfluidic channel.

18. The method of claim 1, wherein the metallic layer extends across a width of the microfluidic channel.

19. The method of claim 1, wherein the metallic layer has a horizontal dimension of greater than 100 microns.

20. The method of claim 1, wherein the metallic layer has a thickness of greater than 10 nanometers.

21. The method of claim 1, wherein the binding and associating steps are performed while a fluid is flowing continuously in the microfluidic channel.

22. The method of claim 1, further comprising quantitatively determining the opacity of the metallic layer.

23. The method of claim 1, wherein the metal colloid associates with the sample component before the sample component binds with the binding partner disposed on the surface of the microfluidic channel.

24. The method of claim 1, wherein the metal colloid associates with the sample component after the sample component binds with the binding partner disposed on the surface of the microfluidic channel.

25. The method of claim 1, wherein the metallic layer has a horizontal dimension of greater than 10 microns.

26. The method of claim 1, wherein the metallic layer is continuous.

27. The method of claim 1, wherein the metallic layer is discontinuous.

28. The method of claim 26, wherein the metallic layer is a web or honeycomb of material that includes passages that allow light to be transmitted therethrough.

29. The method of claim 1, wherein the metallic layer is completely opaque.

30. The method of claim 12, wherein the plurality of surfaces includes at least a first surface including a first binding partner disposed across the first surface and a second surface including a second binding partner disposed across the second surface.

31. The method of claim 30, comprising flowing the metal solution over the first and second surfaces to form a first metallic layer at the first surface and a second metallic layer at the second surface.

32. The method of claim 31, wherein each of the first and second metallic layers has a horizontal dimension of greater than 10 microns.

33. The method of claim 32, wherein the first and second binding partners at the first and second surfaces, respectively, are different from one another.

34. The method of claim 33, wherein each of the first and second binding partners is disposed across a width of the microfluidic channel.

35. The method of claim 9, wherein the metallic layer allows transmittance of less than 50 percent of the light irradiating the metallic layer.

36. The method of claim 1, wherein each of the steps is performed during continuous flow of fluids in the microfluidic channel.

37. The method of claim 1, comprising applying a vacuum to an outlet of the microfluidic channel to induce fluid flow.

38. The method of claim 6, wherein the silver solution comprises silver nitrate.