Electrodes with Multilayer Membranes and Methods of Using and Making the Electrodes

1 - 20 . (canceled)

21 . A membrane structure comprising:

a sensing layer membrane comprising a polymer, an analyte responsive enzyme and a mediator, wherein at least one of the analyte-responsive enzyme and the mediator is chemically bonded to the sensing layer membrane; and

a flux limiting membrane chemically bonded to the sensing layer comprising one or more layers of a polyionic polymer.

22 . The membrane structure of claim 21 , wherein the flux limiting membrane comprises at least one polymer layer selected from the group consisting of a polycationic polymer layer, a polyanionic polymer layer and a zwitterionic polymer layer.

23 . The membrane structure of claim 21 , wherein the flux limiting membrane comprises one or more bilayers, wherein a bilayer comprises a polyanionic polymer layer and a polycationic polymer layer.

24 . The membrane structure of claim 22 , wherein the polycationic polymer comprises one or more nitrogen-containing moieties.

25 . The membrane structure of claim 24 , wherein the one or more nitrogen-containing moieties is pyridine.

26 . The membrane structure of claim 24 , wherein the polycationic polymer comprises quaternized nitrogen.

27 . The membrane structure of claim 22 , wherein the polyanionic polymer comprises one or more acid moieties.

28 . The membrane structure of claim 27 , wherein the polyanionic polymer comprises carboxylate anions.

29 . The membrane structure of claim 27 , wherein the polyanionic polymer comprises sulfonate anions.

30 . The membrane structure of claim 22 , wherein the flux limiting membrane comprises a last layer comprising a polyanionic polymer.

31 . The membrane structure of claim 21 , wherein at least one of the analyte-responsive enzyme and the mediator is covalently bonded to the polymer of the sensing layer membrane.

32 . The membrane structure of claim 31 , wherein at least one of the analyte-responsive enzyme and the mediator forms one or more of carbon-carbon, carbon-nitrogen, or metal-carbon covalent bonds with the polymer of the sensing layer membrane.

33 . The membrane structure of claim 31 , wherein at least one of the analyte-responsive enzyme and the mediator is covalently bonded to the polymer of the sensing layer membrane through a linker moiety.

34 . The membrane structure of claim 31 , wherein the mediator is covalently bonded to the polymer of the sensing layer membrane through one or more ligands of the mediator.

35 . The membrane structure of claim 34 , wherein the one or more ligands comprises a substituted or unsubstituted heterocyclic nitrogen-containing moiety.

36 . The membrane structure of claim 34 , wherein one or more ligands of the mediator are substituents of the polymer backbone of the sensing layer membrane.

37 . The membrane structure of claim 31 , wherein the analyte-responsive enzyme is covalently bonded to the polymer of the sensing layer membrane through one or more side chains of the enzyme.

38 . The membrane structure of claim 37 , wherein the one or more side chains of the analyte-responsive enzyme is selected from the group consisting of substituted or unsubstituted amines, alcohols, thiols, phenols, imidazoles, indols, and carboxylic acids.

39 . The membrane structure of claim 37 , wherein one or more side chains of the analyte-responsive enzyme are substituents of the polymer backbone of the sensing layer membrane.

40 . The membrane structure of claim 21 , wherein the polymer of the sensing layer membrane is crosslinked.

41 . The membrane structure of claim 40 , wherein at least one of the analyte-responsive enzyme and the mediator is crosslinked with the polymer of the sensing layer membrane.

42 . The membrane structure of claim 41 , wherein at least one of the analyte-responsive enzyme and the mediator is crosslinked with the polymer of the sensing layer membrane through one or more crosslinking agents.

43 . The membrane structure of claim 42 , wherein the one or more crosslinking agents is selected from the group consisting of substituted or unsubstituted epoxides, aldehydes, imidoesters, N-hydroxysuccinimide esters, carbodiimides, cyanuric chloride, tetrachlorobenzoquinone, benzoquinone and tetracyanoquinodimethane.

44 . The membrane structure of claim 41 , wherein one or more ligands of the mediator are crosslinkers of the polymer of the sensing layer membrane.

45 . The membrane structure of claim 41 , wherein one or more side chains of the analyte-responsive enzyme are crosslinkers of the polymer of the sensing layer membrane.

46 . The membrane structure of claim 31 , wherein the polymer of the sensing layer membrane forms a coordination complex with the mediator.

47 . The membrane structure of claim 46 , wherein one or more nitrogen-containing moieties of the polymer of the sensing layer membrane forms a coordination complex with a metal of the mediator.

48 . The membrane structure of claim 46 , wherein the mediator is ionically bonded to the polymer of the sensing layer membrane.

49 . The membrane structure of claim 46 , wherein the polymer of the sensing layer membrane comprises at least one positively charged moiety and the mediator comprises at least one negatively charged moiety.

50 . The membrane structure of claim 46 , wherein the polymer of the sensing layer membrane comprises at least one negatively charged moiety and the mediator comprises at least one positively charged moiety.

51 . The membrane structure of claim 21 , wherein the analyte-responsive enzyme is a glucose-responsive enzyme.

52 . The membrane structure of claim 51 , wherein the analyte-responsive enzyme is glucose dehydrogenase (GDH) or glucose oxidase (GOx).

53 . The membrane structure of claim 21 , wherein the analyte-responsive enzyme further comprises an enzyme cofactor.

54 . The membrane structure of claim 53 , wherein the enzyme cofactor is flavin adenine dinucleotide.

55 . The membrane structure of claim 21 , wherein the mediator comprises ruthenium or osmium.