Membrane strip biosensor system for point-of-care testing

• US 7,300,802 B2
• Filed: 04/20/2004
• Issued: 11/27/2007
• Est. Priority Date: 04/25/2003
• Status: Active Grant

• Current Assignee: Biodigit Laboratories Corporation
• Original Assignee: Biodigit Laboratories Corporation
• Inventors: Paek, Se Hwan, Cho, Jung Hwan, Kim, Ser Ka
• Primary Examiner(s): Nguyen; Bao-Thuy L.
• Application Number:
  US10/827,884
• Publication Number:
  US 20040214253A1
• Time in Patent Office: 1,316 Days
• Field of Search: 436/518, 436/514, 436/169, 436/170, 436/808, 436/823, 422 56- 60, 422/82.02, 204/403.01, 204/403.03, 204/403.06, 204/403.05
• US Class Current: 436/514
• CPC Class Codes: C12Q 1/001 Enzyme electrodes
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387
• CPC Class Codes: Y10S 436/808 Automated or kit
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick
• CPC Class Codes: Y10S 436/823 Immunogenic carrier or carrier per se

Abstract

The present invention relates to a biosensor for point-of-care testing (POCT) whose detection sensitivity was remarkably improved by introducing to membrane strip chromatographic assay system a successive cross-flow procedure for immune reaction and enzymatic reaction. The present invention relates to a membrane strip biosensor system that comprises

• (a) a membrane pad (10) for sample application,
• (b) a membrane pad (20) for release of detection binding component, wherein the membrane pad (20) contains label-linked binding component for detection in a dry state,
• (c) a signal generation membrane pad (30) with immobilized binding component for capture,
• (d) a membrane pad (40) for absorption of vertical flow medium,
• (e) a membrane pad (50) for the supply of substrate solution for enzyme,
• (f) a membrane pad (60) for absorption of horizontal flow medium and
• (g) substrate solution,
  wherein the system has a cross-arrangement of two groups of the membrane pads,
  • (I) one group of vertically arranged pads, wherein the pad (10) is partially superimposed and fixed in length at the end of the pad (20), and the pad (20) and the pad (40) are partially superimposed and fixed in length at the both ends of the signal generating membrane pad (30), respectively; and
  • (II) the other group of horizontally arranged pads, wherein the pad (50) and pad (60) are, at the time of signal generation, partially superimposed and fixed at the both lateral sides of the signal generation membrane pad (30), respectively.

Claims

1. A membrane strip biosensor system comprising:

(a) a first group of membrane pads comprising;

(i) a first membrane pad for sample application,

(ii) a second membrane pad for release of a detection binding component, said second membrane pad comprises a tracer-linked binding component specific for and able to bind to an analyte in said sample to form an analyte/binding component complex,

(iii) a third membrane pad that is a signal generation membrane pad with an immobilized binding component specific for and able to capture said analyte/binding component complex,

(iv) a fourth membrane pad for absorption of flow medium;

(b) a second group of membrane pads comprising;

(vi) a fifth membrane pad for the supply of a substrate solution,

(vii) a sixth membrane pad for absorption of excess substrate solution, and

(c) substrate solution;

wherein the system has a cross-arrangement of the two groups of membrane pads;

said first group of membrane pads is arranged in a first flow direction wherein said first membrane pad is partially superimposed and fixed at an end of said second membrane pad, and said second membrane pad and said fourth membrane pad are partially superimposed and fixed one at each end of the third membrane pad, and wherein said sample is applied to said first membrane pad and flows laterally along said second and third membrane pads to said fourth membrane pad to comprise a first flow reaction; and

said second group of membrane pads is arranged in a second flow direction perpendicular to the first flow direction, wherein said fifth and sixth membrane pads are partially superimposed and fixed at both lateral sides of the third membrane pad to form a pathway for a second flow reaction.

2. The membrane strip biosensor system in claim 1, wherein for an electrochemical determination, said signal generation membrane pad further comprises an electrode.

3. The membrane strip biosensor system of claim 1, wherein the system further comprises a container for holding said substrate solution.

4. The membrane strip biosensor system in claim 1, wherein the said first membrane pad for sample application, said second membrane pad for release of detection binding component, and said fifth membrane pad for the supply of substrate solution are glass fiber membranes, the signal generation membrane pad is a nitrocellulose membrane, and said fourth and sixth membrane pads for absorption are a cellulose membranes.

5. The membrane strip biosensor system in claim 1, wherein the tracer-linked binding component for detection comprises either

i) a conjugate of a tracer with a first binding component, or

ii) a first binding component and a conjugate of a tracer with a second binding component specific to the first binding component.

6. The membrane strip biosensor system in claim 5, wherein the binding component for detection is selected from the group consisting of an antibody, an enzyme, a receptor, and DNA that reacts specifically with an analyte in said sample.

7. The membrane strip biosensor system in claim 5, wherein the tracer is an enzyme selected from the group consisting of horseradish peroxidase, alkaline phosphatase, β

-galactosidase, and arthromyces ramosus peroxidase and the substrate solution comprises a chromogenic substrate component specific to said enzyme, and, at the time of signal generation, color change detectable with naked eyes is shown as signal as the result of an enzyme-substrate reaction.

8. The membrane strip biosensor system in claim 5, wherein the tracer is gold colloids and the substrate solution comprises a silver compound, and, at the time of signal generation, color change detectable with naked eyes is shown as signal by chemical catalytic reaction.

9. The membrane strip biosensor system in claim 5, wherein the tracer is horseradish peroxidase or arthromyces ramosus peroxidase and the substrate solution comprises luminol or other luminescent substrate components specific to an enzyme, and at the time of signal generation, a light signal is measured as signal generated by enzyme-substrate reaction.

10. The membrane strip biosensor system in claim 5, wherein the tracer is Co²⁺, Cu²⁺, Mg²⁺, Fe²⁺ or their compounds and the substrate solution comprises luminol or other luminescent substrate component, and at the time of signal generation, a light signal is measured as signal generated by chemical catalytic reaction.

11. The membrane strip biosensor system in claim 5, wherein the tracer is glucose oxidase, urease, penicillin oxidase, or cholesterol oxidase and the substrate solution comprises an electrochemical signal-generating component specific to an enzyme, and, at the time of signal generation, conductivity change, current change, or voltage change is measured as signal generated by enzyme-substrate reaction.

12. The membrane strip biosensor system in claim 1, wherein the binding component for capture is selected from the group consisting of an antibody, an enzyme, a receptor, or a DNA which reacts specifically with said analyte or with said analyte/binding component complex.

13. A biosensor membrane strip system having a plurality of membrane pads in fluid communication, said system comprising:

(a) an immuno-strip comprising

(1) a sample application membrane pad;

(2) a detection reagent-releasing membrane pad positioned to receive sample from said sample application membrane pad, said detection reagent-releasing membrane pad comprising a mobilizable labeled specific binding reagent for binding to an analyte in said sample to form an analyte/detection reagent complex, wherein said mobilizable labeled detection reagent is freely soluble or dispersible in said sample and is free to move with said sample along said detection reagent-releasing membrane pad, whereby it is transported by said sample from said detection reagent-releasing membrane pad to a signal generation membrane pad;

(3) a signal generation membrane pad downstream of said detection reagent-releasing membrane pad, said signal generation membrane pad comprising an immobilized capture reagent for binding to said analyte/detection reagent complex; and

(4) a first absorbent membrane pad downstream of said signal generation membrane pad;

wherein said sample application membrane pad, detection reagent-releasing membrane pad and signal generation membrane pad overlap at their adjacent ends to provide effective contact between the pads thereby forming a first flow path; and

(b) an enzyme substrate solution flow strip comprising;

(5) a substrate solution membrane pad; and

(6) a second absorbent membrane pad,

wherein said substrate solution membrane pad and said second absorbent membrane pads, when brought into fluid communication with said signal generation membrane pad of said immunostrip, form a second flow path that is different from the first flow path.

14. The biosensor membrane strip system of claim 13, wherein said first flow path along said immuno-strip is perpendicular to said second flow path.

15. The biosensor membrane strip system of claim 13, wherein said first fluid flow path is in the same direction as said second fluid flow path.