Materials and methods for detecting oxalate
First Claim
1. A process for preparing an oxalate oxidase composition comprising the steps of preparing an extract from plant tissue, wherein phenylmethylsulfonyl flouride is included during preparation of said extract, precipitating proteins in said extract by bringing said extract to at least about 30% saturation with ammonium sulfate, removing said precipitated proteins and bringing said extract to at least about 60% saturation with ammonium sulfate to precipitate said oxalate oxidase present in said extract, and recovering said oxalate oxidase.
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Abstract
The subject invention pertains to a novel assay device for detecting oxalate in a sample. The assay device comprises enzyme and dye compositions immobilized on a solid carrier matrix. The subject invention can also be used to measure the concentration of oxalate in a sample. The subject invention further pertains to a novel oxalate oxidase composition and methods of preparing the subject enzyme composition. The oxalate oxidase composition can be used in the assay device of the subject invention.
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13 Claims
- 1. A process for preparing an oxalate oxidase composition comprising the steps of preparing an extract from plant tissue, wherein phenylmethylsulfonyl flouride is included during preparation of said extract, precipitating proteins in said extract by bringing said extract to at least about 30% saturation with ammonium sulfate, removing said precipitated proteins and bringing said extract to at least about 60% saturation with ammonium sulfate to precipitate said oxalate oxidase present in said extract, and recovering said oxalate oxidase.
- 8. An oxalate oxidase composition having a specific activity of at least about 10 enzyme units per milligram of protein after ammonium sulfate ((NH4)2 SO4) precipitation.
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9. An oxalate oxidase having a specific activity of at least about 20 enzyme units per milligram of protein after ammonium sulfate ((NH4)2 SO4).
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11. A method for detecting oxalate in a test sample, comprising the steps of:
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(a) contacting said test sample for an effective period of time with an assay device for detecting oxalate, wherein a detectable reaction product is produces when oxalate is present in said test sample, said assay device comprising;
(i) an enzyme composition, wherein oxalate is a substrate for said enzyme composition, and wherein said enzyme composition reacts with oxalate in said sample to produce a first reaction product;
(ii) a dye composition, wherein said dye composition is reactive with said first reaction product to produce a second reaction product, wherein said second reaction product can be detected; and
(iii) a carrier matrix, wherein said enzyme composition and said dye composition are immobilized on said carrier matrix; and(b) detecting said oxalate by detecting the production of said reaction product. - View Dependent Claims (12, 13)
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Specification