Electrochemiluminescent immunoassay and apparatus therefor

• US 4,280,815 A
• Filed: 06/18/1979
• Issued: 07/28/1981
• Est. Priority Date: 06/18/1979
• Status: Expired due to Term

• Current Assignee: Technicon Instruments Corporation
• Original Assignee: Technicon Instruments Corporation
• Inventors: Oberhardt, Bruce J., Wotherspoon, Neil
• Primary Examiner(s): Turk, Arnold
• Application Number:
  US06/049,818
• Time in Patent Office: 771 Days
• Field of Search: 23/230 B, 23/915, 23/927, 424/12, 422/50, 422/55, 422/57, 422/61, 422/68, 204/1 T, 204/1 Y, 204/149
• US Class Current: 436/518
• CPC Class Codes: G01N 21/76 Chemiluminescence; Bioluminescence
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: Y10S 436/805 Optical property
• CPC Class Codes: Y10S 436/806 Electrical property or magnetic property
• CPC Class Codes: Y10S 436/807 Apparatus included in process claim, e.g. physical support structures

Abstract

A method and apparatus for performing a chemiluminescent immunoassay featuring electrochemical techniques to generate an oxidant used to trigger the chemiluminescence of the labelled immunoreactant. The generation of the oxidant is precisely controlled and the oxidant precisely and uniformly delivered to provide a more accurate, precise and sensitive assay.

Claims

1. A method of immunoassaying a sample containing an immunoreactant comprising the steps of:

(a) forming a reaction mixture comprising said sample immunoreactant, a complementary immunoreactant, and a competitive immunoreactant with respect to said sample immunoreactant, said competitive immunoreactant having a chemiluminescent label attached thereto;

(b) confining said reaction mixture between a pair of electrodes;

(c) immobilizing said complementary immunoreactant adjacent one of said electrodes prior to the confining of said reaction mixture;

(d) incubating said reaction mixture to form a reaction product;

(e) applying a controlled voltage across said pair of electrodes for a controlled period of time to generate an oxidant for chemically triggering said chemiluminescently labelled competitive immunoreactant bound to said complementary immunoreactant in said reaction product; and

(f) measuring the chemiluminescence of said bound chemiluminescently labelled competitive immunoreactant.

2. The method of claim 1, wherein said applying step comprises applying said controlled voltage across said pair of electrodes prior to and/or during said incubation step and said measuring step comprises measuring said chemiluminescence during application of said controlled voltage.

3. The method of claim 1, wherein the step of immobilizing comprises immobilizing said complementary immunoreactant on the surface of said one of said electrodes.

4. The method of claim 1, wherein the step of immobilizing comprises immobilizing said complementary immunoreactant on a membrane adjacent said one electrode.

5. The method of claim 1, wherein the step of immobilizing comprises containing said complementary immunoreactant within a gel layer disposed adjacent the surface of said one electrode.

6. The method of claim 1, further comprising the step of:

(g) washing away any excess immunoreactants from between said pair of electrodes after said incubating step and prior to said measuring step.

7. The method of claim 1, further comprising the step of:

(g) releasing said bound chemiluminescently labelled competitive immunoreactant from said immobilized complementary immunoreactant following said measuring step.

8. The method of claim 7, wherein said releasing step comprises changing the pH of the said reaction mixture.

9. The method of claim 7, wherein said releasing step comprises changing the tonicity of said reaction mixture.

10. The method of claim 7, wherein said releasing step comprises changing the temperature of said reaction mixture.

11. The method of claim 7, wherein said releasing step comprises applying a second voltage across said pair of electrodes.

12. The method of claim 7, wherein said releasing step comprises introducing a releasing agent into said mixture.

13. The method of claim 7, wherein said releasing agent is chaotronic.

14. The method of claim 13, wherein said releasing agent comprises a detergent.

15. The method of claim 13, wherein said releasing agent is selected from a group consisting of:

magnesium chloride, a soluble thiocyanate salt and a soluble citrate salt.

16. Apparatus for conducting an immunoassay, comprising:

first and second metallic electrodes defining first and second opposing surfaces, respectively, said first electrode defining an anode and supporting a first immunoreactant on said first surface, and at least one of said electrodes being light transmissive;

means for introducing a reactive solution between said first and second surfaces comprising at least second and third immunoreactants, said second immunoreactant being competitive with said first immunoreactant with respect to binding with said third immunoreactant, said second immunoreactant being labelled with chemiluminescent substance;

means for impressing an electrical potential across said first and second electrodes for generating an oxidant at said first surface to oxidize said chemiluminescent substance; and

means for measuring the chemiluminescence of said chemiluminescent substance, said measuring means comprising a detector means disposed adjacent said one electrode for receiving chemiluminescent radiation generated by oxidation of said chemiluminescent substance.

17. The apparatus of claim 16, wherein said first electrode comprises a membrane to which said first immunoreactant is bound.

18. The apparatus of claim 16, wherein said first electrode comprises a gel layer containing said first immunoreactant.

19. The apparatus of claim 18, wherein said one electrode is integrally formed with said detector means.

20. The apparatus of claim 16, wherein said introducing means comprises a tubular structure having a spiral configuration disposed between said first and second electrodes.

21. The apparatus of claim 20, wherein said tubular structure defines a plurality of reaction compartments disposed between said first and second electrodes.