Compositions and methods for simultaneous inactivation of alkaline phosphatase and peroxidase enzymes during automated multiplex tissue staining assays
AssigneeVentana Medical Systems, Inc.
InventorsJames Grille, Brian D. Kelly, Jerome Kosmeder, II, Eric May, Noemi Sebastiao, Pamela Wirth
Life of this patent
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Abstract
Disclosed are compositions and methods for inactivating one or more enzymes in a biological sample.
Claims25
A method of inactivating one or more enzymes in a biological sample comprising applying between about 50 μL to about 200 μL of an enzyme inactivation composition comprising a polycarboxylic acid, a peroxide, and a preservative to the biological sample, the biological sample comprising one or more reagent or endogenous enzymes, and wherein at least one of the enzyme inactivation composition or the biological sample is maintained at a temperature ranging from between about 25° C. to about 50° C. for a time period ranging from between about 4 minutes to about 16 minutes.
A method of detecting targets in a biological sample, comprising:
- a) contacting the biological sample with a first detection reagent including a first enzyme;
- b) detecting a first signal from the first detection reagent;
- and
- c) inactivating the first enzyme by applying a first enzyme inactivation composition to the biological sample, wherein the first enzyme inactivation composition comprises a polycarboxylic acid having a pH ranging from about 1 to about 3;
- and at least one of a peroxide or a preservative, wherein at least one of the first enzyme inactivation composition or the biological sample is maintained at a first temperature ranging from between about 25° C. to about 50° C. for a first time period ranging from between about 4 minutes to about 16 minutes.
A biological sample comprising one or more enzymes that are either substantially inactivated or completely inactivated, the biological sample prepared by introducing to the biological sample an enzyme inactivation composition comprising a polycarboxylic acid, hydrogen peroxide, and sodium azide, wherein the hydrogen peroxide is present in an amount of about 0.25% to about 5% by total weight of the enzyme inactivation composition, and wherein the sodium azide is present in an amount of about 0.05% to about 1% by total weight of the enzyme inactivation composition, and wherein the enzyme inactivation composition is allowed to remain in contact with the biological sample for at least about 4 minutes at a temperature of between about 25° C. and about 50° C.
A method of detecting one or more targets in a biological sample, comprising:
- a) contacting the biological sample with one or more detection reagents, each detection reagent comprising a different enzyme;
- b) detecting signals from the one or more detection reagents.
- c) inactivating each of the different enzymes of the one or more detection reagents by applying an enzyme inactivation composition to the biological sample, wherein the enzyme inactivation composition comprises a polycarboxylic acid having a pH ranging from about 1 to about 3;
- and at least one of a peroxide or a preservative, wherein at least one of the first enzyme inactivation composition or the biological sample is maintained at a first temperature ranging from between about 25° C. to about 50° C. for a first time period ranging from between about 4 minutes to about 16 minutes.
Dependencies are inferred from the claim text.