Internal calibration system for flow-through assays

• US 20030119204A1
• Filed: 04/25/2002
• Published: 06/26/2003
• Est. Priority Date: 12/24/2001
• Status: Abandoned Application

• Current Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Original Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Inventors: Wei, Ning, Song, Xuedong
• Application Number:
  US10/132,673
• Publication Number:
  US 20030119204A1
• 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 includes two or more calibration regions (e.g., lines, dots, etc.) containing differing amounts of a binder that is configured to bind with the probe conjugates. As a result, calibration signals are generated that can be readily compared (visually, quantitatively, and the like) to a detection signal to determine the presence or quantity of an 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 that contains a binder configured to bind with said probe conjugate, said calibration zone including;

i) a first calibration region containing a first predetermined amount of said binder, said first calibration region being capable of generating a first calibration signal;

ii) a second calibration region containing a second predetermined amount of said binder that is greater than said first predetermined amount of said binder, said second calibration region being capable of generating a second calibration signal, said second calibration signal having a greater intensity than said first calibration signal; and

wherein the relative amount of the analyte within the test sample is determined by comparing said detection signal to said first calibration signal and said second calibration signal.

2. A flow-through assay as defined in claim 1, wherein said detection signal is capable of being visually compared to said first calibration signal and said second calibration signal.

3. A flow-through assay as defined in claim 1, wherein said detection signal is capable of being compared to said first calibration signal and said second calibration signal through the use of an instrument.

4. A flow-through assay as defined in claim 1, wherein a calibration curve is generated by plotting the intensity of said first and second calibration signals versus known levels of the analyte.

5. A flow-through assay as defined in claim 1, wherein said calibration zone further includes a third calibration region containing a third predetermined amount of said binder that is greater than said second predetermined amount of said binder, said third calibration region being capable of generating a third calibration signal that has a greater intensity than said second calibration signal.

6. A flow-through assay as defined in claim 1, wherein said first and said second calibration regions are disposed in a direction that is substantially parallel to the flow of the test sample through said porous membrane.

7. A flow-through assay as defined in claim 1, wherein said binder is a polyelectrolyte.

8. 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.

9. A flow-through assay as defined in claim 8, wherein said detectable probe comprises a latex microparticle.

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

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

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

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

14. 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 a probe conjugate containing a specific binding member and a detectable probe, wherein said porous membrane defines:

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

a calibration zone that contains a binder configured to bind with said probe conjugate, said calibration zone including;

i) a first calibration line containing a first predetermined amount of said binder, said first calibration line being capable of generating a first calibration signal;

ii) a second calibration line containing a second predetermined amount of said binder that is greater than said first predetermined amount of said binder, said second calibration line being capable of generating a second calibration signal, said second calibration signal having a greater intensity than said first calibration signal;

iii) a third calibration line containing a third predetermined amount of said binder that is greater than said second predetermined amount of said binder, said third calibration line being capable of generating a third calibration signal that has a greater intensity than said second calibration signal; and

wherein the relative amount of the analyte within the test sample is determined by comparing said detection signal to said first calibration signal, said second calibration signal, and said third calibration signal.

15. A flow-through assay as defined in claim 14, wherein said detection signal is capable of being visually compared to said first calibration signal, said second calibration signal, and said third calibration signal.

16. A flow-through assay as defined in claim 14, wherein said detection signal is capable of being compared to said first calibration signal, said second calibration signal, and said third calibration signal through the use of an instrument.

17. A flow-through assay as defined in claim 14, wherein a calibration curve is generated by plotting the intensity of said first, second, and third calibration signals versus known amounts of the analyte.

18. A flow-through assay as defined in claim 14, wherein said first, second, and third calibration lines are disposed in a direction that is substantially parallel to the flow of the test sample through said porous membrane.

19. 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:

i) 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;

ii) a calibration zone that contains a binder configured to bind with said uncomplexed probe conjugates, said calibration zone including;

a) a first calibration region containing a first predetermined amount of said binder, said first calibration region being capable of generating a first calibration signal;

b) a second calibration region containing a second predetermined amount of said binder that is greater than said first predetermined amount of said binder, said second calibration region being capable of generating a second calibration signal, said second calibration signal having a greater intensity than said first calibration signal; and

wherein the relative amount of the analyte within the test sample is determined by comparing said detection signal to said first calibration signal and said second calibration signal.

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:

i) 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

ii) a calibration zone that contains a binder configured to bind with said probe conjugates unbound to said capture reagents, said calibration zone including;

a) a first calibration region containing a first predetermined amount of said binder, said first calibration region being capable of generating a first calibration signal;

b) a second calibration region containing a second predetermined amount of said binder that is greater than said first predetermined amount of said binder, said second calibration region being capable of generating a second calibration signal, said second calibration signal having a greater intensity than said first calibration signal; and

wherein the relative amount of the analyte within the test sample is determined by comparing said detection signal to said first calibration signal and said second 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:

i) 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

ii) a calibration zone that contains a binder configured to bind to said probe conjugate/analyte complexes and said probe conjugates remaining unbound to said capture reagents, said calibration zone including;

a) a first calibration region containing a first predetermined amount of said binder, said first calibration region being capable of generating a first calibration signal;

b) a second calibration region containing a second predetermined amount of said binder that is greater than said first predetermined amount of said binder, said second calibration region being capable of generating a second calibration signal, said second calibration signal having a greater intensity than said first calibration signal; and

wherein the relative amount of the analyte within the test sample is determined by comparing said detection signal to said first calibration signal and said second calibration signal.

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