Opposable-element chromatographic assay device for detection of analytes

• US 6,087,184 A
• Filed: 11/10/1997
• Issued: 07/11/2000
• Est. Priority Date: 11/10/1997
• Status: Expired due to Fees

• Current Assignee: Beckman Coulter Incorporated (Danaher Corporation)
• Original Assignee: Beckman Coulter Incorporated (Danaher Corporation)
• Inventors: Magginetti, Paul David, Fitzgerald, Daniel Joseph
• Primary Examiner(s): Chin, Christopher L.
• Application Number:
  US08/967,572
• Time in Patent Office: 974 Days
• Field of Search: 422/55-58, 422/61, 435/7.1, 435/7.5, 435/7.92, 435/7.94, 435/287.1, 435/287.2, 435/287.7, 435/287.9, 435/805, 435/810, 435/962, 435/970, 436/510, 436/512, 436/513, 436/514, 436/518, 436/524, 436/525, 436/528, 436/531, 436/533, 436/534, 436/65, 436/164, 436/169, 436/174, 436/175, 436/177, 436/178, 436/800 , 436/805, 436/810, 436/814, 436/817, 436/818, 436/825
• US Class Current: 436/514
• CPC Class Codes: G01N 30/90 Plate chromatography, e.g. thin layer or paper chromatography
• CPC Class Codes: G01N 33/528 Atypical element structures, e.g. gloves, rods, tampons, toilet paper
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 435/81 Packaged device or kit
• CPC Class Codes: Y10S 435/962 Prevention or removal of interfering materials or reactants or other treatment to enhance results, e.g. determining or preventing nonspecific binding
• CPC Class Codes: Y10S 435/97 Test strip or test slide
• CPC Class Codes: Y10S 436/805 Optical property
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick
• CPC Class Codes: Y10T 436/25 including sample preparation

Abstract

An assay device for detection or determination of an analyte in a sample uses opposable components and is suitable for assay of human chorionic gonadotropin and other protein or glycoprotein hormones. One embodiment of the device comprises: (1) a first opposable component including: (a) a first chromatographic medium having first and second ends and an immobilized first specific binding partner for the analyte in a detection zone; (b) a conjugate pad in operable contact with the first end of the first chromatographic medium, the conjugate pad containing a labeled second specific binding partner for the analyte in resolubilizable form; and (c) a second chromatographic medium having first and second ends and having immobilized thereon in a reference zone a third specific binding partner that specifically binds the labeled second specific binding partner for the analyte and does not bind the analyte, the first end of the second chromatographic medium being in operable contact with the conjugate pad; and (2) a second opposable component including a sample application zone. The assay device can include a timing control to indicate when flow through the chromatographic medium has occurred and the assay can be read, and can also contain a validation zone ensuring that interference from human anti-mouse antibody is not present. Other embodiments of devices are included, as well as methods of use.

Claims

1. A chromatographic assay device for detection or determination of an analyte in a test sample comprising:

(a) a first opposable opponent including;

(i) a first chromatographic medium having first and second ends and an immobilized first specific binding partner for the analyte in a detection zone;

(ii) a conjugate pad in operable contact with the first end of the first chromatographic medium so that fluid applied to the conjugate pad flows from the conjugate pad to the first end of the first chromatographic medium, the conjugate pad including;

(A) a first portion containing a labeled second specific binding partner for the analyte in resolubilizable form;

(B) a second portion; and

(C) a third portion containing a labeled second specific binding partner for the analyte in resolubilizable form, the second portion separating the first portion and the third portion; and

(iii) a second chromatographic medium having first and second ends and having immobilized thereon in a reference zone a third specific binding partner, other than the analyte or an analyte analogue, that specifically binds the labeled second specific binding partner for the analyte and does not bind the analyte, the first end of the second chromatographic medium being in operable contact with the conjugate pad so that fluid applied to the conjugate pad flows from the conjugate pad to the first end of the second chromatographic medium; and

(b) a second opposable component including a sample application zone;

wherein the first and second opposable components are brought into opposition to apply a sample applied to the sample application zone to the conjugate pad to resolubilize the labeled second specific binding partner so that the sample and the resolubilized labeled second specific binding partner in the first portion of the conjugate pad are applied to the first chromatographic medium and the sample and the resolubilized labeled second specific binding partner in the third portion of the conjugate pad are applied to the second chromatographic medium for detection of the analyte.

2. The chromatographic assay device of claim 1 wherein the label is a colloidal particle label.

3. The chromatographic assay device of claim 2 wherein the colloidal particle label is a colloidal carbon label.

4. The chromatographic assay device of claim 1 wherein the label is a visually detectable label.

5. The chromatographic assay device of claim 1 wherein the sample application zone contains at least one reagent for treatment of the sample.

6. The chromatographic assay device of claim 1 wherein the analyte is selected from the group consisting of chorionic gonadotropin (hCG), luteinizing hormone (LH), follicle stimulating hormone (FSH), thyroid stimulating hormone (TSH), corticotropin-releasing hormone (CRH), growth hormone-releasing hormone (GHRH), prolactin, human growth hormone (hGH), β

-lipotropin, corticotropin, β

-endorphin, calcitonin, parathormone, human placental lactogen (hPL), insulin, glucagon, pancreatic polypeptide (PP), secretin, cholecystokinin-pancreozymin, motilin, vasoactive intestinal peptide, gastric inhibitory peptide, and erythropoietin.

7. The chromatographic assay device of claim 6 wherein the analyte is selected from the group consisting of hCG, TSH, LH, and FSH.

8. The chromatographic assay device of claim 7 wherein the analyte is hCG.

9. The chromatographic assay device of claim 8 wherein the first specific binding partner is a monoclonal antibody specific for the a subunit of hCG and the second specific binding partner is a monoclonal antibody specific for the P subunit of hCG and not cross-reactive with luteinizing hormone.

10. The chromatographic assay device of claim 1 where the first opposable component further includes a resolubilizable visible dye, the resolubilizable visible dye being located either in an area between the detection zone and the second end of the first chromatographic medium or in an absorber that is optionally present and, if present, is in operable contact with the second end of the first chromatographic medium to absorb fluid from the first chromatographic medium, and the second opposable component has an aperture for viewing of the visible dye when flow has occurred through the first chromatographic medium and the assay can be read and interpreted.

11. The chromatographic assay device of claim 1 wherein the first opposable component further includes:

(i) a resolubilizable visible dye linked to a first member of an auxiliary specific binding pair that does not bind to any other specific binding partner in the device or to the analyte, the resolubilizable visible dye being located either in an area between the detection zone and the second end of the first chromatographic medium or in an absorber that is optionally present, the absorber, if present, being in operable contact with the second end of the first chromatographic medium to absorb fluid from the second end of the first chromatographic medium and (ii) an immobilized second member of the auxiliary specific binding pair that binds only the first member of the auxiliary specific binding pair in a dye viewing area, and the second opposable component has an aperture for viewing of the visible dye when flow has occurred through the first chromatographic medium and the assay can be read and interpreted.

12. The chromatographic assay device of claim 11 wherein the first member of the auxiliary specific binding pair is biotin and the second member of the auxiliary specific binding pair is avidin or streptavidin.

13. The device of claim 1 wherein the sample application zone contains a fourth specific binding partner that is specific for human antibodies that bind the second specific binding partner and wherein the first chromatographic medium contains, in a validation zone, a fifth immobilized specific binding partner that is specific for human antibodies that bind the second specific binding partner such that the presence of detectable label at the validation zone indicates interference, the validation zone being located closer to the first end of the first chromatographic medium than is the detection zone.

14. The chromatographic assay device of claim 13 wherein the second specific binding partner is a non-human monoclonal antibody, the fourth specific binding partner is specific for human IgG antibodies that bind to the non-human monoclonal antibody, and the fifth immobilized specific binding partner is specific for human IgG antibodies that bind to the non-human monoclonal antibody.

15. The chromatographic assay device of claim 14 wherein the first and second specific binding partners are murine monoclonal antibodies, and wherein the fourth and fifth specific binding partners are each murine IgG or a polymer of murine IgG.

16. The chromatographic assay device of claim 13 wherein the fourth specific binding partner is immobilized in the sample application zone.

17. The chromatographic assay device of claim 13 wherein the fourth specific binding partner is mobile and located at the sample application zone.

18. A method for detecting or determining an analyte in a test sample comprising the steps of:

(a) applying the sample to the sample application zone on the second opposable component of the chromatographic assay device of claim 1;

(b) bringing the first and second opposable components of the chromatographic assay device into operable contact so that the second labeled specific binding partner in the first and third portions of the conjugate pad is resolubilized;

(c) allowing the sample and the resolubilized labeled second specific binding partner in the first portion of the conjugate pad to migrate through the first chromatographic medium and allowing the sample and the resolubilized labeled second specific binding partner in the third portion of the conjugate pad to migrate through the second chromatographic medium; and

(d) observing or measuring the labeled second specific binding partner bound to the first specific binding partner at the detection zone in order to detect or determine the analyte.

19. A chromatographic assay device for detection of an analyte in a sample comprising:

(a) a first opposable component having;

(i) a first chromatographic medium having a first functional zone containing an immobilized first reagent;

(ii) a first conjugate zone in operable contact with the first chromatographic medium so that fluid applied to the first conjugate zone flows to the first chromatographic medium, the first conjugate zone containing a second reagent in resolubilizable form;

(iii) a second chromatographic medium having a second functional zone containing an immobilized third reagent;

(iv) a second conjugate zone in operable contact with the second chromatographic medium so that fluid applied to the second conjugate zone flows to the second chromatographic medium, the second conjugate zone containing a fourth reagent in resolubilizable form; and

(v) an isolation zone that functionally isolates the first conjugate zone and the first chromatographic medium from the second conjugate zone and the second chromatographic medium; and

(b) a second opposable component having a sample application zone for applying the sample,

wherein when the first and second opposable components are brought into opposition, the sample applied to the sample application zone is applied to the first and second conjugate zones and resolubilizes the second and fourth resolubilizable reagents.

20. The chromatographic assay device of claim 19,

wherein the first and second chromatographic media are formed of physically separate pieces of nitrocellulose material, and

wherein the first opposable component comprises a conjugate pad having a first portion constituting the first conjugate zone, a second portion constituting the second conjugate zone, and a third portion between the first and second portion and constituting the isolation zone.

21. The chromatographic assay device of claim 19,

wherein the first and second chromatographic media are formed of a contiguous piece of nitrocellulose material with a separation area between the first and second functional zones, and

wherein the first opposable component comprises a conjugate pad over the separation area, the conjugate pad having a first portion constituting the first conjugate zone, a second portion constituting the second conjugate zone, and a third portion between the first and second portion and constituting the isolation zone.

22. The chromatographic assay device of claim 19, wherein the first and third reagents are unlabeled and the second and fourth reagents are labeled.

23. The chromatographic assay device of claim 22 wherein the label for the second and/or fourth reagent is a colloidal particle label.

24. The chromatographic assay of claim 23 wherein the colloidal particle label is a colloidal carbon label.

25. The chromatographic assay device of claim 22 wherein the label for the second and/or fourth reagent is a visually detectable label.

26. The chromatographic assay device of claim 19, wherein when the sample with the resolubilized second and fourth reagents are applied to the respective first and second chromatographic media, the second and fourth reagents are immobilized by the first and third reagents in the first and second functional zones, respectively.

27. The chromatography assay device of claim 19, wherein the first reagent is an unlabeled first specific binding partner for the analyte, the second reagent is a labeled second specific binding partner for the analyte, the third reagent is a third specific binding partner, other than the analyte or an analyte analogue, that specifically binds the labeled second specific binding partner for the analyte and does not specifically bind the analyte, and the fourth reagent is the labeled second specific binding partner for the analyte.

28. The chromatographic assay device of claim 19, wherein the first reagent is an unlabeled first specific binding partner for the analyte, the second reagent is a labeled second specific binding partner for the analyte, the third reagent is a third specific binding partner, other than the analyte or an analyte analogue, that does not specifically bind the analyte or a specific binding partner that specifically binds the analyte, and the fourth reagent is a labeled fourth specific binding partner that does not specifically bind the analyte or a specific binding partner that specifically binds the analyte, the third and fourth specific binding partners specifically binding each other.

29. The chromatographic assay device of claim 19, wherein the first reagent is an unlabeled first specific binding partner for the analyte, the second reagent is a labeled second specific binding partner for the analyte, the third reagent is the analyte or an analyte analogue, and the fourth reagents is the labeled second specific binding partner for the analyte.

30. The chromatographic assay device of claim 19, wherein the second functional zone contains a predetermined quantity of the third reagent so that a label generated by the fourth reagent in the second functional zone has an intensity equivalent to that of a label generated by the second reagent in the first functional zone when a predetermined concentration of analyte is present in the sample.

31. A method for detecting or determining an analyte in a sample using the chromatographic assay device of claim 30, comprising:

(a) applying the sample to the sample application zone on the second opposable component of the assay device;

(b) bringing the first and second opposable components of the chromatographic assay device into operable contact whereby the second and fourth reagents in the first and second conjugate zones are resolubilized;

(c) allowing the sample and the resolubilized second reagent to migrate through the first chromatographic medium and allowing the sample and the resolubilized fourth reagent to migrate through the second chromatographic medium;

(d) observing or measuring any label in the first functional zone to detect or determine the analyte; and

(e) comparing intensities of labels in the first and second functional zones to determine a relative amount of the analyte present in the sample.

32. The chromatographic assay device of claim 19, wherein the first chromatographic medium further has a dye zone containing a resolubilizable visible dye and a dye viewing area, wherein the first functional zone is located between the dye zone and the first conjugate zone, and the dye zone is located between the dye viewing area and the first functional zone.

33. The chromatographic assay device of claim 32, wherein the first opposable component further has an absorber in operable contact with the first chromatographic medium and located farther away from the first conjugate zone than is the first functional zone so that fluid flown to the first functional zone from the first conjugate zone flows to the absorber, and wherein the dye zone and the dye viewing area are located in the absorber.

34. The chromatographic assay device of claim 32, wherein the resolubilizable visible dye is linked to a first member of an auxiliary specific binding pair that does not specifically bind to any other specific binding partner in the device or to the analyte, and wherein the dye viewing area contains an immobilized second member of the auxiliary specific binding pair that specifically binds only the first member of the auxiliary specific binding pair.

35. The chromatographic assay device of claim 34 wherein the first member of the auxiliary specific binding pair is biotin and the second member of the auxiliary specific binding pair is avidin or streptavidin.

36. A method for detecting or determining an analyte in a sample using the chromatographic assay device of claim 32, comprising:

(a) applying the sample to the sample application zone on the second opposable component of the assay device;

(b) bringing the first and second opposable components of the chromatographic assay device into operable contact whereby the second reagent in the first conjugate zone is resolubilized;

(c) allowing the sample and the resolubilized second reagent to migrate through the first chromatographic medium;

(d) observing any dye in the dye viewing area to determine whether the migration has occurred through the first chromatographic medium; and

(e) observing or measuring any label in the first functional zone to detect or determine the analyte.

37. The chromatographic assay device of claim 19, wherein the second opposable component comprises a backing having first and second designated areas corresponding in position to the first and second functional zones, respectively, so that when the first and second opposable components are brought into opposition, the first and second functional zones are visible through the first and second designated areas, respectively.

38. The chromatographic assay device of claim 19, further comprising an attachment mechanism that attaches the first and second opposable components to each other and allows them to be brought into opposition.

39. The chromatographic assay device of claim 19 wherein the sample application zone contains at least one reagent for treatment of the sample.

40. The chromatographic assay device of claim 19 wherein the analyte is selected from the group consisting of chorionic gonadotropin (hCG), luteinizing hormone (LH), follicle stimulating hormone (FSH), thyroid stimulating hormone (TSH), cotricotropin-releasing hormone (CRH), growth hormone-releasing hormone (GHRH), prolactin, human growth hormone (hGH), β

-lipotropin, corticotropin, β

-endorphin, calcitonin, parathormone, human placental lactogen (hPL), insulin, glucagon, pancreatic polypeptide (PP), secretin, cholecystokinin-pancreozymin, motilin, vasoactive intestinal peptide, gastric inhibitory paptide, and erythropoietin.

41. The chromatographic assay device of claim 40 wherein the analyte is selected from the group consisting of hCG, TSH, LH, and FSH.

42. The chromatographic assay device of claim 41 wherein the analyte is hCG.

43. The chromatographic assay device of claim 19 wherein the first specific binding partner is a monoclonal antibody specific for the a subunit of hCG and the second specific binding partner is a monoclonal antibody specific for the β

subunit of hCG and not cross-reactive with luteinizing hormone.

44. The chromatographic assay device of claim 19 wherein the sample application zone contains a fifth specific binding partner that is specific for human antibodies that bind the second specific binding partner, and wherein the first chromatographic medium further has a validation zone containing a sixth immobilized specific binding partner that is specific for human antibodies that bind the second specific binding partner such that the presence of detectable label at the validation zone indicates interference, the validation zone being located between the first conjugate zone and the first functional zone.

45. The chromatographic assay device of claim 44 wherein the second specific binding partner is a non-human monoclonal antibody, the fifth specific binding partner is specific for human IgG antibodies that bind to the non-human monoclonal antibody, and the sixth specific binding partner is specific for human IgG antibodies that bond to the non-human monoclonal antibody.

46. The chromatographic assay device of claim 44 wherein the fifth specific binding partner is immobilized in the sample application zone.

47. The chromatographic assay device of claim 44 wherein the fifth specific binding partner is mobile and located at the sample application zone.

48. The chromatographic assay device of claim 45 wherein the first and second specific binding partners are murine monoclonal antibodies, and wherein the fifth and sixth specific binding partners are each murine IgG or a derivative or polymer of murine IgG.

49. A method for detecting or determining an analyte in a sample using the chromatographic assay device of claim 44, comprising:

(a) applying the sample to the sample application zone on the second opposable component of the assay device;

(b) bringing the first and second opposable components of the chromatographic assay device into operable contact whereby the second reagent in the first conjugate zone is resolubilized;

(c) allowing the sample and the resolubilized second reagent to migrate through the first chromatographic medium;

(d) observing or measuring any label in the first functional zone to detect or determine the analyte; and

(e) observing any label in the validation zone to determine whether any interference is present.

50. A method for detecting or determining an analyte in a sample using the chromatographic assay device of claim 19, comprising:

(a) applying the sample to the sample application zone on the second opposable component of the assay device;

(b) bringing the first and second opposable components of the chromatographic assay device into operable contact whereby the second and fourth reagents in the first and second conjugate zones are resolubilized;

(c) allowing the sample and the resolubilized second reagent to migrate through the first chromatographic medium and allowing the sample and the resolubilized fourth reagent to migrate through the second chromatographic medium;

(d) observing or measuring any label in the first functional zone to detect or determine the analyte; and

(e) observing or measuring any label in the second functional zone to determine whether the assay has run properly.