Polyelectrolytic internal calibration system of a flow-through assay

• US 20030124739A1
• Filed: 04/25/2002
• Published: 07/03/2003
• Est. Priority Date: 12/24/2001
• Status: Active Grant

• Current Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Original Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Inventors: Wei, Ning, Song, Xuedong, Sayre, Curt
• Granted Patent: US 7,651,841 B2
• Time in Patent Office: Days
• US Class Current: 436/514
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387

Abstract

A flow-through assay for detecting the quantity of an analyte residing in a test sample is provided. The flow-through assay contains a porous membrane that is in fluid communication with probe conjugates that contain a specific binding member and a detectable probe. The porous membrane also defines a detection zone and a calibration zone. The calibration zone contains a polyelectrolyte substantially non-diffusively immobilized on the porous membrane. The polyelectrolyte is capable of generating a detectable calibration signal that can be readily compared (visually, quantitatively, and the like) to a detection signal to determine the amount of analyte in the test sample.

Claims

1. A flow-through assay for detecting the presence or quantity of an analyte residing in a test sample, said flow-through assay comprising a porous membrane, wherein said porous membrane is in fluid communication with a probe conjugate that contains a specific binding member and a detectable probe, said porous membrane defining:

a detection zone that contains a capture reagent that is capable of binding to the analyte or the probe conjugate, wherein said detection zone is capable of generating a detection signal that represents the presence or absence of the analyte;

a calibration zone, wherein a polyelectrolyte is substantially non-diffusively immobilized on said porous membrane within said calibration zone, said polyelectrolyte being capable of binding to said probe conjugate to generate a calibration signal such that the relative amount of the analyte within the test sample is determined by comparing said detection signal to said calibration signal.

2. A flow-through assay as defined in claim 1, wherein said polyelectrolyte has a net positive charge.

3. A flow-through assay as defined in claim 2, wherein said polyelectrolyte is selected from the group consisting of polylysine, polyethylenimine, epichlorohydrin-functionalized polyamines or polyamidoamines, polydiallyidimethyl-ammonium chloride, cationic cellulose derivatives, and combinations thereof.

4. A flow-through assay as defined in claim 1, wherein said polyelectrolyte has a net negative charge.

5. A flow-through assay as defined in claim 1, wherein said polyelectrolyte is amphiphilic.

6. A flow-through assay as defined in claim 1, wherein said calibration zone contains at least two distinct regions having a different concentration of said polyelectrolyte.

7. A flow-through assay as defined in claim 1, wherein said polyelectrolyte is ionically bonded to a functional group present on the surface of said porous membrane.

8. A flow-through assay as defined in claim 1, wherein said polyelectrolyte is covalently bonded to a functional group present on the surface of said porous membrane.

9. A flow-through assay as defined in claim 8, wherein said polyelectrolyte is crosslinked to said functional group.

10. A flow-through assay as defined in claim 1, wherein said detectable probe is selected from the group consisting of chromogens, catalysts, fluorescent compounds, chemiluminescent compounds, radioactive labels, direct visual labels, liposomes, and combinations thereof.

11. A flow-through assay as defined in claim 1, wherein said specific binding member is selected from the group consisting of antigens, haptens, antibodies, and complexes thereof.

12. A flow-through assay as defined in claim 1, wherein said capture reagent is selected from the group consisting of antigens, haptens, antibodies, and complexes thereof.

13. A flow-through assay as defined in claim 1, wherein the assay is a sandwich-type assay.

14. A flow-through assay as defined in claim 1, wherein the assay is a competitive-type assay.

15. A flow-through assay for detecting the presence or quantity of an analyte residing in a test sample, said flow-through assay comprising a porous membrane in fluid communication with probe conjugates containing a specific binding member and a detectable probe, said probe conjugates being configured to combine with the analyte in the test sample when contacted therewith such that probe conjugate/analyte complexes and uncomplexed probe conjugates are formed, wherein said porous membrane defines:

a detection zone in which a capture reagent is substantially non-diffusively immobilized on said porous membrane, said capture reagent being capable of binding to said probe conjugate/analyte complexes, wherein said detection zone is capable of generating a detection signal;

a calibration zone in which a predetermined amount of polyelectrolyte is substantially non-diffusively immobilized on said porous membrane, said polyelectrolyte being capable of binding to said uncomplexed probe conjugates to generate a calibration signal such that the relative amount of the analyte within the test sample is determined by comparing said detection signal to said calibration signal.

16. A flow-through assay as defined in claim 15, wherein said calibration zone contains at least two distinct regions having a different concentration of said polyelectrolyte.

17. A flow-through assay as defined in claim 15, wherein said polyelectrolyte is ionically bonded to a functional group present on the surface of said porous membrane.

18. A flow-through assay as defined in claim 15, wherein said polyelectrolyte is covalently bonded to a functional group present on the surface of said porous membrane.

19. A flow-through assay as defined in claim 18, wherein said polyelectrolyte is crosslinked to said functional group.

20. A flow-through assay for detecting the presence or quantity of an analyte residing in a test sample, said flow-through assay comprising a porous membrane in fluid communication with probe conjugates that contain a specific binding member and a detectable probe, wherein said porous membrane defines:

a detection zone in which a predetermined amount of capture reagent is substantially non-diffusively immobilized on said porous membrane, said capture reagent being capable of binding to said probe conjugates and to the analyte, wherein said detection zone is capable of generating a detection signal, and

a calibration zone in which a polyelectrolyte is substantially non-diffusively immobilized on said porous membrane, said polyelectrolyte being capable of binding said probe conjugates unbound to said capture reagents to generate a calibration signal such that the relative amount of the analyte within the test sample is determined by comparing said detection signal to said calibration signal.

21. A flow-through assay as defined in claim 20, wherein said specific binding member is identical to the analyte.

22. A flow-through assay for detecting the presence or quantity of an analyte residing in a test sample, said flow-through assay comprising a porous membrane in fluid communication with probe conjugates that contain a specific binding member and a detectable probe, said probe conjugates being configured to combine with the analyte in the test sample when contacted therewith such that probe conjugate/analyte complexes and uncomplexed probe conjugates are formed, wherein said porous membrane defines:

a detection zone in which a capture reagent is substantially non-diffusively immobilized on said porous membrane, said capture reagent being capable of binding to said uncomplexed probe conjugates, wherein said detection zone is capable of generating a detection signal, and

a calibration zone in which a polyelectrolyte is substantially non-diffusively immobilized on said porous membrane, said polyelectrolyte being capable of binding to said probe conjugate/analyte complexes and said probe conjugates remaining unbound to said capture reagents, said calibration zone being capable of generating a calibration signal such that the relative amount of the analyte within the test sample is determined by comparing said detection signal to said calibration signal

23. A flow-through assay as defined in claim 22, wherein said capture reagent is identical to the analyte.