Distal tip of biosensor transducer comprising enzyme for deamination

• US 8,323,956 B2
• Filed: 01/22/2009
• Issued: 12/04/2012
• Est. Priority Date: 06/01/2001
• Status: Expired due to Term

• Current Assignee: Colorado State University Research Foundation, Regents of The University of Minnesota (University of Minnesota)
• Original Assignee: Colorado State University Research Foundation
• Inventors: Reardon, Kenneth F., Wackett, Lawrence Philip
• Primary Examiner(s): NAFF, DAVID M
• Application Number:
  US12/358,140
• Publication Number:
  US 20090221014A1
• Time in Patent Office: 1,412 Days
• Field of Search: None
• US Class Current: 435/287.1
• CPC Class Codes: C12Q 1/002 Electrode membranes
• CPC Class Codes: G01N 2333/978 acting on carbon to nitrogen bonds other than peptide bonds (3.5)
• CPC Class Codes: G01N 2333/988 Lyases (4.), e.g. aldolases, heparinase, enolases, fumarase

Abstract

Enzymatic biosensors and methods of producing distal tips for biosensor transducers for use in detecting one or more analytes selected from organic compounds susceptible to dehalogenation, organic compounds susceptible to oxygenation, organic compounds susceptible to deamination, organosulfate compounds susceptible to hydrolysis, and organophosphate compounds susceptible to hydrolysis are disclosed herein, as well as biosensor arrays, methods of detecting and quantifying analytes within a mixture, and devices and methods for delivering reagents to enzymes disposed within the distal tip of a biosensor.

Claims

1. A distal tip of a biosensor transducer for use in detecting an organic compound susceptible to deamination, the tip comprising:

a biocomponent comprising one or more enzymes for carrying out deamination of the organic compound, said biocomponent immobilized to a surface of the tip by one or more of (a) entrapment within a hydrogel;

(b) entrapment within a polymeric network;

(c) encapsulation;

(d) covalent bonding; and

(e) adsorption;

wherein said biocomponent has been treated to maintain a period of enzymatic efficacy; and

wherein said biocomponent is stabilized to said tip by one or more of crosslinking a surface of said immobilized biocomponent, crosslinking a polymer layer to said biocomponent, adding a gel-hardening agent to said biocomponent, adding a stabilizing agent to said biocomponent, and modifying a component used to immobilize said biocomponent to said surface of the tip.

2. The distal tip of claim 1, wherein the biosensor transducer is selected from the group consisting of an optical transducer, a pH optode, a pH electrode, a field-effect transistor (FET) and an ion-selective electrode (ISE).

3. The distal tip of claim 1, the one or more enzymes selected from hydrolases of subclass EC 3.5 and lyases of subclass EC 4.3.

4. The distal tip of claim 1, wherein said biocomponent is treated according to a treatment selected from the group consisting of applying an inhibitor of protein synthesis, adding a protease inhibitor, freeze drying and dry heating.

5. The distal tip of claim 1, wherein said biocomponent is selected from whole isolated cells, purified enzymes and combinations thereof.

6. The distal tip of claim 1, wherein the organic compound is selected from the group consisting of melamine, ammeline, ammelide, serine, biotin and aniline.

7. The distal tip of claim 1, said biocomponent stabilized to said tip by crosslinking the polymer layer, wherein said crosslinked polymer layer is selected from the group consisting of poly-L-lysine, polyethylenimine, polyacrylic acid, polyvinyl alcohol, polyacrylamide and polyurethane.

8. A distal tip of a biosensor transducer for use in detecting an organic compound susceptible to deamination, the tip comprising:

a biocomponent comprising at least one enzyme selected from the group consisting of hydrolases from EC family 3.5 and lyases from EC family 4.3 for carrying out deamination of the organic compound, said biocomponent immobilized to a surface of the tip by one or more of (a) entrapment within a hydrogel;

(b) entrapment within a polymeric network;

(c) encapsulation;

(d) covalent-bonding; and

(e) adsorption,

wherein said biocomponent is stabilized to said tip by one or more of crosslinking a surface of said immobilized biocomponent, crosslinking a polymer layer to said biocomponent, adding a gel-hardening agent to said biocomponent, adding a stabilizing agent to said biocomponent, and modifying a component used to immobilize said biocomponent to said surface of the tip.

9. The distal tip of claim 8, wherein the biosensor transducer is selected from the group consisting of an optical transducer, a pH optode, a pH electrode, a field-effect transistor (FET) and an ion-selective electrode (ISE).

10. The distal tip of claim 8, wherein said biocomponent is treated according to a treatment selected from the group consisting of applying an inhibitor of protein synthesis, adding a protease inhibitor, freeze drying and dry heating.

11. The distal tip of claim 8, wherein said biocomponent is selected from whole isolated cells, purified enzymes and combinations thereof.

12. The distal tip of claim 8, wherein the organic compound is selected from the group consisting of melamine, ammeline, ammelide, serine, biotin and aniline.

13. The distal tip of claim 8, said biocomponent stabilized to said tip by crosslinking the polymer layer, wherein said crosslinked polymer layer is selected from the group consisting of poly-L-lysine, polyethylenimine, polyacrylic acid, polyvinyl alcohol, polyacrylamide and polyurethane.

14. A distal tip of a biosensor transducer for use in detecting organic compounds, the tip comprising:

a biocomponent comprising at least one enzyme selected from the group consisting of hydrolases from EC family 3.5 and lyases from EC family 4.3 for carrying out deamination of one organic compound; and

one or more of (i) an oxygenase from EC family 1.13 for carrying out an oxidation of one organic compound, (ii) an oxygenase from EC family 1.14 for carrying out an oxidation of one organic compound, (iii) a dehalogenase for carrying out dehalogenation of one organic compound, and (iv) a hydrolase from subclass EC 3.1 for carrying out hydrolysis of one organic compound, said biocomponent immobilized to a surface of the tip by one or more of (a) entrapment within a hydrogel;

(b) entrapment within a polymeric network;

(c) encapsulation;

(d) covalent-bonding; and

(e) adsorption,

wherein said biocomponent is stabilized to said tip by one or more of crosslinking a surface of said immobilized biocomponent, crosslinking a polymer layer to said biocomponent, adding a gel-hardening agent to said biocomponent, adding a stabilizing agent to said biocomponent, and modifying a component used to immobilize said biocomponent to said surface of the tip.

15. The distal tip of claim 14, further comprising a device for delivering one or more reagents.

16. The distal tip of claim 15, wherein the reagent is selected from oxygen and hydrogen peroxide.

17. The distal tip of claim 14, said biocomponent stabilized to said tip by crosslinking the polymer layer, wherein said crosslinked polymer layer is selected from the group consisting of poly-L-lysine, polyethylenimine, polyacrylic acid, polyvinyl alcohol, polyacrylamide and polyurethane.

18. The distal tip of claim 14, wherein the biosensor transducer is selected from the group consisting of an optical transducer, a pH optode, a pH electrode, a field-effect transistor (FET), an ion-selective electrode (ISE), and an oxygen optode.

19. The distal tip of claim 14, wherein the organic compound is selected from the group consisting of melamine, ammeline, ammelide, serine, biotin and aniline.

20. The distal tip of claim 14, wherein said biocomponent is treated according to a treatment selected from the group consisting of applying an inhibitor of protein synthesis, adding a protease inhibitor, freeze drying and dry heating.