Reduction of the hook effect in membrane-based assay devices

• US 7,662,643 B2
• Filed: 07/20/2007
• Issued: 02/16/2010
• Est. Priority Date: 12/19/2002
• Status: Expired due to Fees

• Current Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Original Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Inventors: Wei, Ning, Huang, Yanbin
• Primary Examiner(s): NGUYEN, BAO THUY L
• Application Number:
  US11/780,671
• Publication Number:
  US 20080014659A1
• Time in Patent Office: 942 Days
• Field of Search: 435/7.1, 435/7.92, 435/288.6, 435/287.1, 436/514, 436/518, 436/523, 436/805, 436/809, 422 56- 61
• US Class Current: 436/514
• CPC Class Codes: G01N 33/538 by sorbent column, particles or resin strip , i.e. sorbent materials
• CPC Class Codes: G01N 33/5432 Liposomes or microcapsules
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387
• CPC Class Codes: Y10S 436/805 Optical property
• CPC Class Codes: Y10S 436/809 Multifield plates or multicontainer arrays

Abstract

A membrane-based assay device for detecting the presence or quantity of an analyte residing in a test sample is provided. The device utilizes a chromatographic zone on which is disposed a plurality of microporous particles. The chromatographic zone can effectively reduce the “hook effect” in a simple, efficient, and relatively inexpensive manner. In particular, the plurality of microporous particles allows larger-sized analyte/probe complexes to reach the detection zone before the uncomplexed analyte. Because the uncomplexed analyte is substantially inhibited from competing with the complexes for the binding sites at the detection zone, the incidence of “false negatives” may be limited, even at relatively high analyte concentrations.

Claims

1. A method for detecting the presence or quantity of an analyte residing in a test sample, said method comprising:

i) providing a flow-through assay device comprising a porous membrane, said porous membrane being in communication with conjugated detection probes capable of generating a detection signal, said porous membrane defining a chromatographic zone within which a plurality of particles having micropores are immobilized and a detection zone located downstream from said chromatographic zone, wherein a capture reagent is immobilized within said detection zone;

ii) contacting a test sample containing the analyte with said conjugated detection probes so that analyte/probe complexes and uncomplexed analyte are formed;

iii) allowing said analyte/probe complexes and said uncomplexed analyte to reach said chromatographic zone and then said detection zone, wherein said analyte/probe complexes move through said chromatographic zone at a faster rate than said uncomplexed analyte such that said analyte/probe complexes reach said detection zone before said uncomplexed analyte; and

iv) detecting said detection signal generated by said detection probes within said detection zone.

2. The method of claim 1, wherein said particles define a plurality of spaces therebetween, said spaces having an average size that is greater than the average size of the micropores of said particles.

3. The method of claim 2, wherein the average size of said micropores is at least about 100% less than the average size of said spaces.

4. The method of claim 2, wherein the average size of said micropores is at least about 150% less than the average size of said spaces.

5. The method of claim 2, wherein the average size of said micropores is at least about 100% less than the average size of said spaces.

6. The method of claim 1, wherein said particles are formed from polystyrenes, polyacrylamides, polyacrylonitriles, silica beads, or combinations thereof.

7. The method of claim 1, wherein the surface of said particles is chemically inert to the analyte.

8. The method of claim 1, further comprising measuring the intensity of the detection signal generated within said detection zone.

9. The method of claim 1, wherein said porous membrane further comprises a calibration zone a capable of generating a calibration signal, wherein the amount of the analyte within the test sample is determined from said detection signal as calibrated by said calibration signal.

10. The method of claim 9, wherein said porous membrane is in communication with calibration probes, said calibration probes generating said calibration signal when present within said calibration zone.

11. The method of claim 10, further comprising generating a calibration curve by plotting the intensity of the detection signal calibrated by the intensity of the calibration signal for a plurality of predetermined analyte concentrations.

12. The method of claim 1, wherein said particles have an average diameter of from about 0.1 to about 100 microns.

13. The method of claim 1, wherein said particles have an average diameter of from about 1 to about 10 microns.

14. The method of claim 1, wherein said porous membrane contains pores having an average size of greater than about 200 nanometers.

15. The method of claim 1, wherein said porous membrane contains pores having an average size of from about 200 to about 5000 nanometers.

16. The method of claim 1, wherein said porous membrane contains pores having an average size of from about 200 to about 2500 nanometers.

17. The method of claim 1, wherein the detections probes are conjugated with an antibody.

18. The method of claim 1, wherein the capture reagent is an antibody.

19. The method of claim 1, wherein the micropores have an average size less than about 100 nanometers.

20. The method of claim 1, wherein the micropores have an average size from about 5 to about 100 nanometers.

21. The method of claim 1, wherein the micropores have an average size from about 10 to about 60 nanometers.