Interdigitated electrode arrays for liposome-enhanced immunoassay and test device

• US 5,958,791 A
• Filed: 09/27/1996
• Issued: 09/28/1999
• Est. Priority Date: 09/27/1996
• Status: Expired due to Fees

• Current Assignee: Cornell Research Foundation Incorporated (Cornell University), Rheonix, Inc.
• Original Assignee: Innovative Biotechnologies, Inc.
• Inventors: Durst, Richard Allen, Roberts, Matthew A, Rule, Geoffrey S., Montagna, Richard A.
• Primary Examiner(s): Chin, Christopher L.
• Assistant Examiner(s): NGUYEN, BAO THUY L
• Application Number:
  US08/722,901
• Time in Patent Office: 1,096 Days
• Field of Search: 422/56, 422/57, 422/70, 422/82.01, 422/82.03, 422/98, 422/99, 422/110, 427/213.3, 427/213.34, 427/213.35, 435/3, 435/7.1, 435/7.93, 435/287.7, 435/970, 436/516, 436/518, 436/530, 436/541, 204/182.7, 204/286, 204/288, 204/290 R, 204/297 R
• US Class Current: 436/514
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2565/501 being an array of oligonucleotides
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: G01N 33/586 Liposomes, microcapsules or cells
• CPC Class Codes: Y10S 435/97 Test strip or test slide

Abstract

A test device for detecting or determining an analyte in a test solution includes an absorbent material having contact, liposome lysing and electrochemical measurement portions. The contact portion is positioned for contact with and uptake of the test solution. The liposome lysing portion is segregated from the contact portion and has a liposome lysing agent bound thereto. The liposome lysing portion is further either positioned between the contact portion and the electrochemical measurement portion, or partially or completely coincides with the electrochemical measurement portion. The electrochemical measurement portion comprises a first conductor comprising a plurality of fingers disposed on the absorbent material, and a second conductor similarly comprising a plurality of fingers disposed on the absorbent material, where the fingers of the first and second conductors are interdigitated to form an array.

Claims

1. A test device for detecting or quantifying an analyte in a test sample, said test device comprising an absorbent material, said absorbent material comprising:

a contact portion at or proximate to a first end of said absorbent material;

an electrochemical measurement portion at a location on said absorbent material which is positioned away from the first end, wherein said electrochemical measurement portion comprises an electrochemical detector cell consisting of a first conductor comprising a plurality of fingers disposed on said absorbent material and a second conductor comprising a plurality of fingers disposed on said absorbent material, wherein said fingers of said first conductor are interdigitated with said fingers of said second conductor, and wherein said conductors induce redox cycling; and

a liposome lysing portion segregated from said contact portion and having a liposome lysing agent bound thereto, wherein said liposome lysing portion either is positioned between said contact portion and said electrochemical measurement portion, or partially or completely coincides with said electrochemical measurement portion.

2. A test device according to claim 1, wherein said absorbent material further comprises a competitive binding portion positioned between and segregated from said contact portion and said liposome lysing portion and having a binding material for the analyte bound to said competitive binding portion.

3. A test device according to claim 2, further comprising a reference electrode disposed on said absorbent material between said competitive binding portion and said electrochemical measurement portion.

4. A test device according to claim 3, wherein said reference electrode is electrically connected to said first conductor and said second conductor.

5. A test device according to claim 2, wherein said binding material is present in said competitive binding portion in a concentration of at least 1 μ

g/cm² absorbent material.

6. A test device according to claim 1 or 2, wherein said first conductor and said second conductor are electrically connected to one another.

7. A test device according to claim 1 or 2, further comprising a support material on which said absorbent material is mounted.

8. A test device according to claim 1 or 2, wherein said absorbent material is nitrocellulose.

9. A test device according to claim 8, wherein said nitrocellulose has a pore size of from 0.05 μ

m to about 50 μ

m.

10. A test device according to claim 1 or 2, wherein said contact portion further comprises a wicking portion at said first end.

11. A test device according to claim 1 or 2, wherein said absorbent material has been treated with one or more blocking agents, surfactants, or mixtures thereof.

12. A test device according to claim 11, wherein said blocking agents are selected from the group consisting of proteinaceous blocking reagents which inhibit binding of molecules having a molecular weight of greater than about 1000 with said absorbent material and polymer blocking reagents which inhibit binding of molecules having a molecular weight of less than about 1000 with said absorbent material.

13. A test device according to claim 1, further comprising a reference electrode disposed on said absorbent material between said contact portion and said electrochemical measurement portion.

14. A test device according to claim 13, wherein said reference electrode is electrically connected to said first conductor and said second conductor.

15. A test device according to claim 13 or 3, wherein said reference electrode comprises silver, lead or mixtures thereof.

16. A test device according to claim 1 or 2, wherein either or both of said first conductor and said second conductor comprise one or more materials selected from the group consisting of platinum, gold, graphite, and silver.

17. A test device according to claim 1 or 2, wherein each of said first conductor and said second conductor comprise from 2 to 500 fingers.

18. A test device according to claim 1 or 2, wherein said fingers of said first and second conductors are each from about 1 μ

m to about 20 μ

m wide and are spaced from about 0.5 μ

m to about 10 μ

m apart.

19. A method for detecting or quantifying an analyte in a test sample, comprising:

providing a test device comprising an absorbent material, said absorbent material comprising;

a contact portion at or proximate to a first end of said absorbent material;

an electrochemical measurement portion at a location on said absorbent material which is positioned away from the first end, wherein said electrochemical measurement portion comprises an electrochemical detector cell consisting of a first conductor comprising a plurality of fingers disposed on said absorbent material and a second conductor comprising a plurality of fingers disposed on said absorbent material, wherein said fingers of said first conductor are interdigitated with said fingers of said second conductor and wherein said first conductor and said second conductor are electrically connected to one another; and

a liposome lysing portion segregated from said contact portion and having a liposome lysing agent bound thereto, wherein said liposome lysing portion either is positioned between said contact portion and said electrochemical measurement portion, or partially or completely coincides with said electrochemical measurement portion;

combining a binding material specific for the analyte with a conjugate of an analyte analog and liposomes and the test sample in an electrolyte mixture, wherein said liposomes comprise an electroactive marker;

incubating the mixture for a time sufficient to permit competition between any analyte present in the test sample and the conjugate for the binding material;

contacting the mixture with said contact portion of said absorbent material after said incubating;

applying a potential to said conductors, wherein said potential is sufficient to induce redox cycling of said marker;

allowing the mixture to migrate from said contact portion through said electrochemical measurement portion of said absorbent material after said incubating, wherein migration of aggregates of conjugate and binding material formed during said incubating is inhibited by said absorbent material, and whereby said liposomes are lysed by said liposome lysing agent to release said marker, and said marker undergoes redox cycling induced by said conductors causing current to flow between said first and second conductors;

detecting the presence or amount of said current; and

correlating the presence or amount of said current with the presence or amount, respectively, of the analyte in the test sample.

20. A method according to claim 19, wherein the analyte analog is either the analyte or a reactive analog of the analyte.

21. A method according to claim 19, wherein said contacting is carried out by inserting the contact portion of the absorbent material into the mixture.

22. A method according to claim 19, wherein the contact portion further comprises a wicking portion at the first end, said contacting being carried out by spotting the mixture onto the absorbent material in the contact portion outside of the wicking zone, and said allowing comprises inserting the wicking portion into a wicking reagent.

23. A method according to claim 19, wherein said correlating is used to determine the amount of analyte in the test sample and is carried out by comparing the amount of the current flowing between the first and second conductors with one or more reference standards having known concentrations of the analyte for particular current amounts to determine the analyte concentration in the test sample relative to the known concentrations.

24. A method according to claim 19, wherein the liposomes are prepared from one or more phospholipids, glycolipids, steroids, alkyl phosphates, or fatty acid esters.

25. A method according to claim 19, wherein the analyte is an antigen or hapten, and the binding material is an antibody recognizing the antigen or the hapten.

26. A method according to claim 19, wherein said absorbent material further comprises a region for accumulation of aggregates formed from said conjugate and said binding material, wherein said region for accumulation is positioned away from said liposome lysing portion and either between said liposome lysing portion and said contact portion, or in said contact portion.

27. A method according to claim 19, wherein said test device further comprises a reference electrode disposed on said absorbent material between said contact portion and said electrochemical measurement portion, and wherein said reference electrode is electrically connected to said first conductor and said second conductor.

28. A method according to claim 27, wherein said reference electrode comprises silver, lead or mixtures thereof.

29. A method according to claim 19, wherein either or both of said first conductor and said second conductor comprise one or more materials selected from the group consisting of platinum, gold, graphite, and silver.

30. A method according to claim 19, wherein each of said first conductor and said second conductor comprise from 2 to 500 fingers.

31. A method according to claim 19, wherein said fingers of said first and second conductors are each from about 1 μ

m to about 20 μ

m wide and are spaced from about 0.5 μ

m to about 10 μ

m apart.

32. A method for detecting or quantifying an analyte in a test sample, comprising:

providing a test device comprising an absorbent material, said absorbent material comprising;

a contact portion at or proximate to a first end of said absorbent material;

an electrochemical measurement portion at a location on said absorbent material which is positioned away from the first end, wherein said electrochemical measurement portion comprises an electrochemical detector cell consisting of a first conductor comprising a plurality of fingers disposed on said absorbent material and a second conductor comprising a plurality of fingers disposed on said absorbent material, wherein said fingers of said first conductor are interdigitated with said fingers of said second conductor and wherein said first conductor and said second conductor are electrically connected to one another;

a liposome lysing portion segregated from said contact portion and having a liposome lysing agent bound thereto, wherein said liposome lysing portion either is positioned between said contact portion and said electrochemical measurement portion, or partially or completely coincides with said electrochemical measurement portion; and

a competitive binding portion positioned between and segregated from said contact and said liposome lysing portions on said absorbent material and having a binding material for the analyte bound to said competitive binding portion;

contacting an electrolytic mixture of the test sample and a conjugate of an analyte analog and liposomes with said contact portion of said absorbent material, wherein said liposomes comprise an electroactive marker;

applying a potential to said conductors, wherein said potential is sufficient to induce redox cycling of said marker;

allowing the mixture to migrate from said contact portion through said electrochemical measurement portion of said absorbent material, whereby said liposomes are lysed by said liposome lysing agent to release said marker, and said marker undergoes redox cycling induced by said conductors causing current to flow between said first and second conductors;

detecting the presence or amount of said current; and

correlating the presence or amount of said current with the presence or amount, respectively, of the analyte in the test sample.

33. A method according to claim 32, wherein the analyte analog is either the analyte or a reactive analog of the analyte.

34. A method according to claim 32, wherein said contacting is carried out by inserting the contact portion of said absorbent material into said mixture.

35. A method according to claim 32, wherein said contact portion further comprises a wicking portion at the first end, said contacting is carried out by spotting said mixture onto said absorbent material in said contact portion outside of said wicking zone, and said allowing comprises inserting said wicking portion into a wicking reagent.

36. A method according to claim 32, wherein said correlating is used to determine the amount of analyte in the test sample and is carried out by comparing the amount of the current flowing between the first and second conductors with one or more reference standards having known concentrations of the analyte for particular current amounts to determine the analyte concentration in the test sample relative to the known concentrations.

37. A method according to claim 32, wherein said liposomes are prepared from one or more phospholipids, glycolipids, steroids, alkyl phosphates, or fatty acid esters.

38. A method according to claim 32, wherein said analyte is an antigen or hapten, and said binding material is an antibody for said antigen or said hapten.

39. A method according to claim 32, wherein said test device further comprises a reference electrode disposed on said absorbent material between said contact portion and said electrochemical measurement portion, and wherein said reference electrode is electrically connected to said first conductor and said second conductor.

40. A method according to claim 39, wherein said reference electrode comprises silver, lead or mixtures thereof.

41. A method according to claim 32, wherein either or both of said first conductor and said second conductor comprise one or more materials selected from the group consisting of platinum, gold, graphite, and silver.

42. A method according to claim 32, wherein each of said first conductor and said second conductor comprise from 2 to 500 fingers.

43. A method according to claim 32, wherein said fingers of said first and second conductors are each from about 1 μ

m to about 20 μ

m wide and are spaced from about 0.5 μ

m to about 10 μ

m apart.