Porous strip form assay device method

• US 5,141,850 A
• Filed: 02/07/1990
• Issued: 08/25/1992
• Est. Priority Date: 02/07/1990
• Status: Expired due to Term

• Current Assignee: Church & Dwight Company Incorporated
• Original Assignee: Hygeia Sciences, Inc. (The Procter & Gamble Company)
• Inventors: Sigillo, Eric C., Cicia, Nancy J., MacDonnell, Paul C., Cole, Francis X.
• Primary Examiner(s): Kepplinger, Esther L.
• Assistant Examiner(s): Spiegel, Carol A.
• Application Number:
  US07/475,486
• Time in Patent Office: 930 Days
• Field of Search: 435/7.5, 435/7.92, 435/7.94, 435/969, 435/970, 435/971, 436/525, 436/535, 436/538, 436/540-541, 436/810, 436/818, 422/56, 422/58
• US Class Current: 436/525
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387
• CPC Class Codes: Y10S 435/969 Multiple layering of reactants
• CPC Class Codes: Y10S 435/97 Test strip or test slide
• CPC Class Codes: Y10S 435/971 Capture of complex after antigen-antibody reaction
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick
• CPC Class Codes: Y10S 436/818 Human chorionic gonadotropin

Abstract

An immunoassay method comprising applying an aqueous solution containing the analyte antigen to one end of a multi-zoned test strip device such that the solution moves along the strip by capillary action. The zones are arranged so that the solution (a) first contacts and reconstitutes dry, diffusible labelled component comprising colloidal gold conjugated to an antibody specific for said analyte antigen and then (b) contacts and reconstitutes dry, diffusible biotinylated second antibody specific for said analyte antigen such that a diffusible, dispersed sandwich reaction product forms. The reaction product diffuses along the strip with the solution and into a zone containing capture component consisting of a latex and avidin complex which avidin collects the reaction product by means of reaction with its biotin moiety. Thus, gold particles are collected and concentrated in the detection zone for visual determination.

Claims

1. An immunoassay of an aqueous sample for determining an immunoreactive analyte, said process comprising the steps of:

providing an immunoassay device which comprises (1) an element being formed from a bibulous carrier material having a porous structure through which an aqueous dispersion system is capable of diffusing by capillary action, and (2) a capture component localized at a detection zone in the strip element, said capture component comprising a capturing species capable of specifically binding to a capturable species, said detection zone being spaced from at least one of the ends of the strip element;

forming an aqueous dispersion system containing (a) the aqueous sample to be assayed for said analyte, (b) a first batch of dispersed first particles comprising a first coupling product of a first immunoreactive substance and a detectable label, and (c) a second batch of dispersed second particles comprising a second coupling product of a second immunoreactive substance and the capturable species, said first and said second immunoreactive substances being the same or different and being capable of specifically binding either competitively or non-competitively to said analyte or to one another as a function of the presence or quantity of said analyte in said system to thereby form a dispersed composite reaction product comprising both first and second particles;

causing said aqueous dispersion system to diffuse along said strip element and through the bibulous material thereof and into said detection zone so as to bring the composite reaction product into contact with said capture component to thereby immobilize and concentrate the composite reaction product at the detection zone; and

evaluating the detection zone for the presence of the detectable label as an indicator of the presence or quantity of the immunoreactive analyte in the aqueous sample.

2. The process as set forth in claim 1, wherein the first and second particles are initially both contained in said bibulous carrier material in a dry, reconstitutable, water-dispersible form, and wherein said aqueous dispersion system is formed by contacting the dry, reconstitutable particles with said aqueous sample.

3. The process as set forth in claim 2, wherein the contact between the aqueous sample and the dry, reconstitutable particles is caused to occur by first brining the strip element into contact with the aqueous sample and then allowing the sample to diffuse through the bibulous carrier material and into contact with the dry, reconstitutable particles.

4. The process as set forth in claim 3, wherein said dry, reconstitutable, water-dispersible particles are initially contained in one or more contact zones in said bibulous carrier material, and said aqueous dispersion system is formed by diffusion of said aqueous sample through said contact zones.

5. The process as set forth in claim 4, wherein said first batch of particles is contained in a first contact zone in said bibulous material and said second batch of particles is contained in a second contact zone in said bibulous material.

6. The process as set forth in claim 5, wherein said step of bringing the strip element into contact with the aqueous sample comprises immersing said one end of the base element in the aqueous sample.

7. The process as set forth in claim 1, wherein said detectable label comprises a metal-containing third particle.

8. The process as set forth in claim 7, wherein said first coupling product is prepared by coupling the first immunoreactive substance directly to the metal-containing third particle.

9. The process as set forth in claim 7, wherein said metal containing third particles are metal sol particles.

10. The process as set forth in claim 9, wherein said metal sol particles have a particle size in the range of from about 25 to about 1000 Angstroms.

11. The process as set forth in claim 9, wherein said metal sol particles are gold sol particles.

12. The process as set forth in claim 1, wherein one of said capturing and capturable species comprises biotin and the other species comprises avidin.

13. The process as set forth in claim 12, wherein said capturable species comprises biotin and said capturing species comprises avidin.

14. The process as set forth in claim 1, wherein said capture component comprises a third coupling product of a fourth particle and said capturing species, said fourth particle being deposited and thus immobilized in the pores of the immobilized carrier material at said detection zone.

15. The process as set forth in claim 14, wherein said capturable species comprises biotin and said capturing species comprises avidin.

16. The process as set forth in claim 5, wherein said first and second contact zones are spaced apart longitudinally of said strip element.

17. The process as set forth in claim 5, wherein said first and second contact zones are disposed between said one end of the strip element and said detection zone in longitudinally spaced relationship relative to the detection zone.

18. The process as set forth in claim 5, wherein said detectable label comprises a metal-containing third particle.

19. The process as set forth in claim 18, wherein said first coupling product is prepared by coupling the first immunoreactive substance directly to the metal-containing third particle.

20. The process as set forth in claim 18, wherein said metal containing third particles are metal sol particles.

21. The process as set forth in claim 20, wherein said metal sol particles have a particle size in the range of from about 25 to about 1000 Angstroms.

22. The process as set forth in claim 20, wherein said metal sol particles are gold sol particles.

23. The process as set forth in claim 5, wherein said capture component comprises a third coupling product of a fourth particle and said capturing species, said fourth particle being deposited and thus immobilized in the pores of said bibulous carrier material at said detection zone.

24. The process as set forth in claim 23, wherein said capturable species comprises biotin and said capturing species comprises avidin.

25. The process as set forth in claim 5, wherein said first and second immunoreactive substances each specifically binds a respective different site of the analyte to form a sandwich.

26. The process as set forth in claim 25, wherein said first and second immunoreactive substances are each antibodies and the analyte is an antigen which is specifically bound by both antibodies.

27. The process as set forth in claim 5, wherein said first and second immunoreactive substances specifically bind each other.

28. The process as set forth in claim 27, wherein one of said immunoreactive substances is an antibody and the other is an antigen which is specifically bound by said antibody.

29. The process as set forth in claim 27, wherein the first immunoreactive substance is an antibody and the second immunoreactive substance is an antigen which is specifically bound by said antibody.

30. A kit of materials for immunoassay of an aqueous sample for determining an immunoreactive analyte, said kit comprising:

an immunoassay device which comprises (1) an elongated strip element having a pair of opposite ends, said strip element being formed from a bibulous carrier material having a porous structure through which an aqueous dispersion system is capable of diffusing by capillary action, and (2) a capture component localized at a detection zone in the base element, said capture component comprising a capturing species capable of specifically binding to a capturable species, said detection zone being spaced from at least one of the ends of the strip element;

a first bath of dry, reconstitutable, water-dispersible first particles comprising a first coupling product of a first immunoreactive substance and a detectable label; and

a second batch of dry, reconstitutable, water-dispersible second particles comprising a second coupling product of a second immunoreactive substance and said capturable species,

the first and second particles of said first and second batches thereof being dispersible upon coming into contact with said aqueous sample to thereby form an aqueous dispersion system which contains said sample and dispersed first and second particles, said first and second immunoreactive substances being the same or different and being capable of specifically binding either competitively or non-competitively to sad analyte or to one another as a function of the presence or quantity of said analyte in said aqueous dispersion system to thereby form a dispersible composite reaction product comprising both first and second particles,

said bibulous carrier material being such that the aqueous dispersion system will diffuse along the strip element and through the bibulous material so as to bring the composite reaction product into contact with the capture component localized at said detection zone to thereby immobilize and concentrate the composite reaction product at the detection zone so that the detection zone may then be evaluated for the presence of the detectable label as an indicator of the presence or quantity of the immunoreactive analyte in the aqueous sample.

31. The kit as set forth in claim 30, wherein said first and second particles are contained in the bibulous carrier material.

32. The kit as set forth in claim 31, wherein said first and second particles are contained in one or more contact zones in said bibulous material.

33. The kit as set forth in claim 32, wherein said first batch of particles is contained in a first contact zone in said bibulous material and said second batch of particles is contained in a second contact zone in said bibulous material.

34. The kit as set forth in claim 33, wherein said first and second contact zones are spaced apart longitudinally on or in said strip element.

35. The kit as set forth in claim 33, wherein said first and second contact zones are disposed between said one end of the strip element and said detection zone in longitudinally spaced relationship relative to the detection zone.

36. The kit as set forth in claim 34, wherein said first and second contact zones are disposed between said one end of the strip element and said detection zone in longitudinally spaced relationship relative to the detection zone.

37. The kit as set forth in claim 36, wherein said detectable label comprises a metal-containing third particle.

38. The kit as set forth in claim 37, wherein said first coupling product is prepared by coupling the first immunoreactive substance directly to the metal-containing third particle.

39. The kit as set forth in claim 37, wherein said metal containing third particles are metal sol particles.

40. The kit as set forth in claim 39, wherein said metal sol particles have a particle size in the range of from about 25 to about 1000 Angstroms.

41. The kit as set forth in claim 39, wherein said particles are gold sol particles.

42. The kit as set forth in claim 36, wherein one of said capturable and capturing species comprises biotin and other species comprises avidin.

43. The kit as set forth in claim 42, wherein said capturable species comprises biotin and said capturing species comprises avidin.

44. The kit as set forth in claim 36, wherein said capture component comprises a third coupling product of a fourth particle and said capturing species, said fourth particle being deposited and thus immobilized in the pores of said bibulous carrier material at said detection zone.

45. The kit as set forth in claim 44, wherein said capturable species comprises biotin and said capturing species comprises avidin.

46. The kit as set forth in claim 36, wherein said first and second immunoreactive substances each specifically binds a respective different site of the analyte to form a sandwich.

47. The kit as set forth in claim 46, wherein said first and second immunoreactive substances are each antibodies and the analyte is an antigen which is specifically bound by both antibodies.

48. The kit as set forth in claim 36, wherein said first and second immunoreactive substances specifically bind each other.

49. The kit as set forth in claim 48, wherein one of said immunoreactive substances is an antibody and the other is an antigen which is specifically bound by said antibody.

50. The kit as set forth in claim 48, wherein the first immunoreactive substance is an antibody and the second immunoreactive substance is an antigen which is specifically bound by said antibody.

51. The kit as set forth in claim 30, wherein is included a wick member disposed in intimate contact with said element at said one end thereof.

52. The kit as set forth in claim 36, wherein is included a wick member disposed in intimate contact with said element at said one end thereof.