Interrupted-flow assay device

• US 5,468,648 A
• Filed: 12/07/1993
• Issued: 11/21/1995
• Est. Priority Date: 05/29/1991
• Status: Expired due to Term

• Current Assignee: Smithkline Diagnostics, Inc. (GlaxoSmithKline PLC)
• Original Assignee: Smithkline Diagnostics, Inc. (GlaxoSmithKline PLC)
• Inventors: Chandler, Howard M.
• Primary Examiner(s): Spiegel, Carol A.
• Application Number:
  US08/163,341
• Time in Patent Office: 714 Days
• Field of Search: 422/56-60, 435/7.1, 435/5, 435/6, 435/7.92-7.95, 435/970, 435/973, 435/974, 436/514, 436/518, 436/530, 436/525, 436/538, 436/540, 436/807, 436/810
• US Class Current: 436/518
• CPC Class Codes: B01D 15/3804 Affinity chromatography
• CPC Class Codes: B01L 2300/0825 Test strips
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 3/5023 with a sample being transported to, and subsequently stored in an absorbent for analysis
• CPC Class Codes: B01L 3/5055 Hinged, e.g. opposable surfaces
• CPC Class Codes: G01N 30/02 Column chromatography
• CPC Class Codes: G01N 30/30 of temperature
• CPC Class Codes: G01N 30/463 for multidimensional chromatography
• CPC Class Codes: G01N 30/90 Plate chromatography, e.g. thin layer or paper chromatography
• CPC Class Codes: G01N 33/5302 Apparatus specially adapted for immunological test procedures
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/54386 Analytical elements
• 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 435/97 Test strip or test slide
• CPC Class Codes: Y10S 435/973 Simultaneous determination of more than one analyte
• CPC Class Codes: Y10S 435/974 Aids related test
• CPC Class Codes: Y10S 436/807 Apparatus included in process claim, e.g. physical support structures
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick

Abstract

The present invention provides chromatographic assay devices that can perform multiple assays simultaneously in the same test strip, as well as methods for their use. One of the assays can be an immunological assay to detect an antigen, such as human chorionic gonadotropin, while another assay can be a serological assay to detect an antibody, such as anti-rubella antibody. An assay device according to the present invention can comprise: (1) a first opposable component including at least one chromatographic medium having a specific binding partner to the first analyte and a specific binding partner to the second analyte immobilized thereto in separate, discrete, non-overlapping zones; and (2) a second opposable component including an absorber. The first and second opposable components are configured such that bringing the first and second opposable components into opposition causes the absorber to come into operable contact with at least one chromatographic medium so that the zone containing the specific binding partner to the first analyte is functionally divided from the zone containing the specific binding partner to the second analyte so that both analytes can be detected.

Claims

1. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising at least one chromatographic medium having (1) a first end comprising a conjugate zone impregnated with a labeled specific binding partner to the first analyte, (2) an intermediate first zone comprising an immobilized specific binding partner to the first analyte, and (3) a second end having a second zone comprising an immobilized antigen which specifically binds to the second analyte;

(b) a second opposable component having an absorber which is brought into fluid communication with an area of the at least one chromatographic medium between the first and second zones thereof when the first and second opposable components are brought into physical contact; and

(c) a labeled specific binding partner to the second analyte impregnated in either the first or the second opposable component in a zone in downstream fluid communication with the second zone;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively.

2. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising at least one chromatographic medium having (1) a first end, (2) an intermediate first zone comprising an immobilized specific binding partner to the first analyte, and (3) a second end having a second zone comprising an immobilized antigen which specifically binds to the second analyte;

(b) a second opposable component having (1) a first applicator impregnated with a labeled specific binding partner for the first analyte which is brought into fluid communication with the first zone when the first and second opposable components are brought into physical contact, (2) an absorber which is brought into fluid communication with an area of the at least one chromatographic medium between the first and second zones thereof when the first and second opposable components are brought into physical contact; and

(3) a second applicator impregnated with a labeled specific binding partner for the second analyte which is brought into fluid communication with the second zone, when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively.

3. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising at least one chromatographic medium having (1) a first end comprising a first applicator impregnated with a labeled specific binding partner to the first analyte, (2) an intermediate first zone comprising an immobilized specific binding partner to the first analyte, and (3) a second end having a second zone comprising an immobilized antigen which specifically binds to the second analyte;

(b) a second opposable component having (1) an absorber which is brought into fluid communication with an area of the at least one chromatographic medium between the first and second zones thereof when the first and second opposable components are brought into physical contact, and (2) a second applicator impregnated with a labeled specific binding partner for the second analyte which is brought into fluid communication with the second zone when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively; and

,

wherein the absorber substantially excludes the labeled specific binding partner to the second analyte from the first zone by blocking the labeled specific binding partner to the second analyte from migrating into the first zone.

4. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising

(i) at least one chromatographic medium having (1) a first end, (2) an intermediate first zone comprising an immobilized specific binding partner to the first analyte, and (3) a second end having a second zone comprising an immobilized antigen which specifically binds to the second analyte, and

(ii) a sample preparation zone located between the first zone and the second zone; and

,

(b) a second opposable component having (1) a first applicator impregnated with a labeled specific binding partner for the first analyte which is brought into fluid communication with the first zone when the first and second opposable components are brought into physical contact, (2) an absorber which is brought into fluid communication with an area of the at least one chromatographic medium between the first and second zones thereof when the first and second opposable components are brought into physical contact; and

(3) a second applicator impregnated with a labeled specific binding partner for the second analyte which is brought into fluid communication with the second zone, when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively; and

,

wherein physical contacting of the absorber divides the first zone from the second zone so that the labeled specific binding partner to the second analyte is substantially excluded from the first zone by blocking the labeled specific binding partner to the second analyte from flowing into the first zone.

5. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising

(i) at least one chromatographic medium having in order of fluid communication (1) a first end, (2) a first applicator, (3) a conjugate zone impregnated with a labeled specific binding partner to the first analyte bridging the first end and the first applicator, (4) a first zone comprising an immobilized specific binding partner to the first analyte, (5) a second zone comprising an immobilized antigen which specifically binds to the second analyte, (6) a fluid conductor, and (7) a second end;

(b) a second opposable component comprising

(i) an absorber which is brought into fluid communication with an area of the at least one chromatographic medium between the first and second zones thereof when the first and second opposable components are brought into physical contact; and

(ii) a second applicator impregnated with a labeled specific binding partner for the second analyte which is brought into fluid communication with the fluid conductor, when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively.

6. The assay device of claim 5 wherein the second analyte is an antibody produced by a mammalian species in response to an antigen, wherein the antigen is the same as the immobilized antigen, and the labeled specific binding partner is a labeled antibody that specifically binds to the second analyte by binding to the constant region the second analyte.

7. The assay device of claim 6 wherein the labeled antibody binds the second analyte by binding to a species-specific determinant.

8. The assay device of claim 5 wherein the first and second zones of the at least one chromatographic medium are of differing porosities.

9. The assay device of claim 5 wherein the at least one chromatographic medium is nitrocellulose.

10. A method for determining the presence or amount of at least two analytes in an aqueous sample, comprising the sequential steps of:

(a) applying a first aliquot of the sample to the first applicator of the assay device of claim 5;

(b) allowing the sample to migrate from the first applicator through the conjugate zone and then through at least the fluid conductor of the at least one chromatographic medium so that a first ternary complex is formed in the first zone if the first analyte is present in the sample, the first ternary complex comprising the first analyte, the labeled specific binding partner to the first analyte, and the immobilized specific binding partner to the first analyte;

(c) bringing the first and second opposable components into physical contact to cause the second applicator to come into fluid communication with the at least one chromatographic medium to draw fluid from the second applicator through the fluid conductor and through the second zone of the at least one chromatographic medium from the fluid conductor to the absorber so that a second ternary complex is formed in the second zone if the second analyte is present in the sample, the second ternary complex comprising the second analyte, the labeled specific binding partner to the second analyte, and the immobilized antigen to the second analyte; and

,

(d) determining the presence or amount of the at least two analytes in the aqueous sample by observing and/or measuring any detectable signal produced by the labeled specific binding partner to at first analyte bound in the first ternary complex in the first zone and any detectable signal produced by the labeled specific binding partner to the second analyte bound in the second ternary complex in the second zone.

11. The method of claim 10 wherein the labeled specific binding partner for the first analyte and the labeled specific binding partner to the second analyte each is labeled with a visually detectable label and the step of observing and/or measuring the detectable signals is performed visually.

12. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising

(i) at least one chromatographic medium having in order of fluid communication (1) a first end, (2) a first fluid conductor, (3) a sample preparation zone, (4) a first zone comprising an immobilized specific binding partner to the first analyte, (5) a second zone comprising an immobilized antigen which specifically binds to the second analyte, (6) a second fluid conductor, and (7) a second end;

(b) a second opposable component comprising

(i) a first applicator impregnated with a resolubilizable labeled specific binding partner for the first analyte which is brought into fluid communication with the first fluid conductor when the first and second opposable components are brought into physical contact,

(ii) an absorber which is brought into fluid communication with the sample preparation zone of the at least one chromatographic medium when the first and second opposable components are brought into physical contact, and

(iii) a second applicator impregnated with a resolubilizable labeled specific binding partner for the first analyte which is brought into fluid communication with the first fluid conductor, when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively.

13. The assay device of claim 12 wherein the second analyte is an antibody produced by a mammalian species in response to an antigen, wherein the antigen is the same as the immobilized antigen, and the labeled specific binding partner is a labeled antibody that specifically binds to the second analyte by binding to the constant region the second analyte.

14. The assay device of claim 13 wherein the labeled antibody binds the second analyte by binding to a species-specific determinant.

15. The assay device of claim 12 wherein the first and second zones of the at least one chromatographic medium are of different porosities.

16. The assay device of claim 15 wherein the chromatographic medium is nitrocellulose.

17. A method for determining the presence or amount of at least two analytes in an aqueous sample, comprising the sequential steps of:

(a) applying a first aqueous liquid to the second applicator of the assay device of claim 12 to resolubilized the labeled specific binding partner to the second analyte;

(b) applying a first aliquot of the sample to the first applicator to resolubilize the labeled specific binding partner to the first analyte;

(c) applying a second aliquot of the sample to the sample preparation zone;

(d) allowing the second aliquot of the sample applied to the sample preparation zone to migrate through at least the second fluid conductor of at least one chromatographic medium;

(e) bringing the first and second opposable components into physical contact to cause the absorber to come into contact with the at least one chromatographic medium, the first applicator to come into contact with the first fluid conductor, and the second applicator to come into contact with the second fluid conductor;

(f) allowing the first aliquot of the sample and the resolubilized labeled specific binding partner to the first analyte to migrate through at least the first zone of the at least one chromatographic medium so that a first ternary complex is formed in the first zone if the first analyte is present in the sample, the first ternary complex comprising the first analyte, the labeled specific binding partner to the first analyte, and the immobilized specific binding partner to the first analyte; and

, allowing the resolubilized labeled specific binding partner to the second analyte to migrate through at least the second zone of the at least one chromatographic medium so that a second ternary complex is formed in the second zone of the second analyte is present in the sample, the second ternary complex comprising the second analyte, the labeled specific binding partner to the second analyte, and the immobilized antigen to the second analyte; and

(g) determining the presence or amount of the first analyte and the second analyte in the aqueous sample by observing and/or measuring any detectable signal produced by the labeled specific binding partner to the first analyte bound in the first ternary complex in the first zone and any detectable signal produced by the labeled specific binding partner to the second analyte bound in the second ternary complex in the second zone.

18. The method of claim 17 wherein the labeled specific binding partner for the first analyte and the labeled specific binding partner to the second analyte are each labeled with a visually detectable label and the step of observing and/or measuring the detectable signals is performed visually.

19. An assay device for detecting at least two analytes in an aqueous sample, a first analyte that is an antigen and a second analyte that is an antibody, the assay device comprising:

(a) a first opposable component comprising

(i) a first chromatographic medium having in order of fluid communication (1) a first end, (2) a first fluid conductor, (3) a first zone comprising an immobilized specific binding partner to the first analyte, and (4) a second end;

(ii) a second chromatographic medium having in order of fluid communication (1) a first end, (2) a sample preparation zone, (3) a second zone comprising an immobilized antigen which specifically binds to the second analyte, (4) a second fluid conductor, and (5) a second end;

(b) a second opposable component comprising

(i) a first applicator impregnated with a resolubilizable labeled specific binding partner for the first analyte which is brought into fluid communication with the first fluid conductor when the first and second opposable components are brought into physical contact,

(ii) an absorber which is brought into fluid communication with the sample preparation zone of the at least one chromatographic medium when the first and second opposable components are brought into physical contact, and

(iii) a second applicator impregnated with a resolubilizable labeled specific binding partner for the first analyte which is brought into fluid communication with the first fluid conductor, when the first and second opposable components are brought into physical contact;

wherein the first and second opposable components are capable of being brought into physical contact to provide a unidirectional chromatographic specific binding assay for the first analyte and a bidirectional chromatographic specific binding assay for the second analyte by physically contacting the absorber to the first opposable component after migration of an applied aqueous sample from the first end to the second end of the first opposable component in order to reverse fluid flow direction between the first and second ends, and detecting the first and second analytes by detecting formation of labeled ternary complexes in the first and second zones, respectively.

20. The assay device of claim 19 wherein the second analyte is an antibody produced by a mammalian species in response to an antigen, wherein the antigen is the same as the immobilized antigen, and the labeled specific binding partner is a labeled antibody that specifically binds to the second analyte by binding to the constant region the second analyte.

21. The assay device of claim 20 wherein the labeled antibody binds the second analyte by binding to a species-specific determinant.

22. The assay device of claim 19 wherein the first and second chromatographic media are of differing porosities.

23. The assay device of claim 22 wherein the first and second chromatographic media are each nitrocellulose.

24. A method for determining the presence or amount of at least two analytes simultaneously in an aqueous sample, comprising the sequential steps of:

(a) applying a first aqueous liquid to the second applicator of the assay device of claim 19 to resolubilize the labeled specific binding partner to the second analyte;

(b) applying a first aliquot of the sample to the first applicator of the assay device to resolubilize the labeled specific binding partner to the first analyte;

(c) applying a second aliquot of the sample to the sample preparation zone;

(d) allowing the second aliquot of the sample applied to the sample preparation zone to migrate through at least the second fluid conductor of the second chromatographic medium;

(e) bringing the first and second opposable components into physical contact to cause the absorber to come into contact with the sample preparation zone and the first chromatographic medium, to cause the first applicator to come into contact with the first fluid conductor, and to cause the second applicator to come into contact with the second fluid conductor;

(f) allowing the first aliquot of the sample and the resolubilized labeled specific binding partner to the first analyte to migrate through at least the first zone of the first chromatographic medium and allowing the resolubilized labeled specific binding partner to the second analyte to migrate through at least the second zone of the second chromatographic medium so that a first ternary complex is formed in the first zone of the first chromatographic medium if the first analyte is present in the sample, the first ternary complex comprising the first analyte, the labeled specific binding partner to the first analyte, and the immobilized specific binding partner to the first analyte; and

, so that a second ternary complex is formed in second zone of the second chromatographic medium if the second analyte is present in the sample, the second ternary complex comprising the second analyte, the labeled specific binding partner to the second analyte, and the immobilized antigen to the second analyte; and

(g) determining the presence or amount of the first analyte and the second analyte in the aqueous sample by observing and/or measuring any detectable signal produced by the labeled specific binding partner to the first analyte bound in the first ternary complex in the first zone in the first chromatographic medium and any detectable signal produced by the labeled specific binding partner to the second analyte bound in the second ternary complex in the second zone of the chromatographic medium.

25. The method of claim 24 wherein the labeled specific binding partner for the first analyte and the labeled specific binding partner to the second analyte are each labeled with a visually detectable label and the step of observing and/or measuring the detectable signals is performed visually.