Device and method for the quantitative determination of heparin in mammalian blood plasma
AssigneeMiles Laboratories, Inc.
InventorsRonald G. Sommer, Alfred C. Greenquist
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Abstract
A device for the quantitative determination of heparin in mammalian blood plasma, a method for the preparation of such device and a method for determining the plasma's heparin concentration using the device. The device, which is prepared using a freeze-drying technique, comprises a first upper layer of a carrier matrix material containing a buffer, AT-III and thrombin and a second, lower layer of a carrier matrix containing a thrombin sensitive fluorogenic or chromogenic substrate being capable of interacting with thrombin in such a manner that a time-related chemical change detectable by fluorometric or spectrophotometric means takes place when thrombin and the substrate are contacted in a suitable liquid environment.
Claims15
A method for the preparation of a three layer device for the quantitative determination of heparin in mammalian blood plasma which comprises the steps of:
- (a) providing a first strip of carrier matrix material having incorporated therewith a buffer and a thrombin sensitive fluorogenic or chromogenic substrate, said substrate being capable of interacting with thrombin in such a manner that a time related chemical change detectable by fluorometric or spectrophotometric means takes place when thrombin and the substrate are contacted in a suitable liquid environment;
- (b) contacting a second strip of a carrier matrix material with a fresh, aqueous solution containing a buffer, AT-III and thrombin which solution has been maintained at a temperature below about 20.degree. C. since before addition of the thrombin thereto to saturate the second strip with the solution;
- (c) removing the second strip from the solution, quick freezing it, and eliminating solvent by freeze-drying to provide a second strip containing buffer, AT-III and thrombin;
- (d) attaching the first strip to a strip of water impermeable material;
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- (e) attaching the second strip to the exposed side of the first strip to form a three-layered device suitable for the intended purpose.
A three layer test device suitable for the quantitative determination of heparin in mammalian blood plasma which comprises:
- i. a first upper layer of a carrier matrix material containing a buffer, AT-III and thrombin;
- ii. a second layer adjacent to and in contact with the first layer said second layer comprising a carrier matrix containing a thrombin sensitive fluorogenic or chromogenic substrate and a buffer said substrate being capable of interacting with thrombin in such a manner that a time-related chemical change detectable by fluorometric or spectrophotometric means takes place when thrombin and the substrate are contacted in a suitable liquid environment;
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- iii. a third layer of a water impermeable material beneath the second layer.
Dependencies are inferred from the claim text.