EP0505604A1
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Highly purified thrombin preparation
Assignee: Warner Lambert Co LLC
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Filing: 1991-03-28
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Granted: —
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Expires: 2011-03-28
Abstract
Abstract An ultra-pure, clear thrombin solution having a high specific activity is described as well as a method of manufacture.
Inventors
- 👤 Amal Boctor
- 👤 Surendra Mehta
- 👤 Galen Radebaugh
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-24 23:38
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/14; C12N9/48; C12N9/50; C12N9/64; C12N9/6421; C12N9/6424; C12N9/6429; C12Y; C12Y304/00; C12Y304/21; C12Y304/21005; A; A61; A61K; A61K38/00
Claims Analysis
Total Claims: 13
Claim 1
Independent Claim
6 words
An ultra-pure, clear, colorless thrombin solution.
Claim 2
Dependent Claim
19 words
An ultra-pure, clear, colorless thrombin solution having a specific activity of from 4000 to 11,000 Units per mg protein.
Claim 3
Dependent Claim
25 words
An ultra-pure, clear, colorless thrombin solution having a specific activity of from 4000 to 11,000 Units per mg protein as determined by a modified NIH-method.
Claim 4
Dependent Claim
19 words
The thrombin solution of Claim 3 having a specific activity of from 4000 to 11,000 Units per mg protein.
Claim 5
Dependent Claim
64 words
An ultra-pure, clear thrombin solution prepared by reacting prothrombin with purified thromboplastin and treating the resulting thrombin after centrifugation by eluting the supernatant through an anion-exchange agarose column; freezing then thawing up to about 25°C the desired eluant fractions; centrifuging, and eluting the supernatant through a cation-exchange agarose column with a 0.1 M to 1 M sodium chloride salt gradient in a phosphate buffer.
Claim 6
Dependent Claim
25 words
The thrombin solution of Claim 5 having a specific activity of from 4000 to 11,000 Units per mg protein as determined by a modified NIH-method.
Claim 7
Dependent Claim
19 words
The thrombin solution of Claim 6 having a specific activity of from 4000 to 11,000 Units per mg protein.
Claim 8
Dependent Claim
74 words
A process for preparing an ultra-pure, clear thrombin solution which comprises (a) reacting prothrombin with purified thromboplastin; centrifuging the resulting protein suspension; (b) eluting the supernatant through an anion-exchange agarose column; freezing then thawing up to about 25°C the desired eluant fractions; (c) centrifuging, and eluting the supernatant through a cation-exchange agarose column with a 0.1 M to 1 M sodium chloride salt gradient in a phosphate buffer, and collecting the desired eluant fractions.
Claim 9
Dependent Claim
74 words
A process as claimed in Claim 8, wherein the purified thromboplastin is prepared by (a) homogenizing bovine/veal lung tissue with 25 mM phosphate buffer, pH 6.5, containing about 0.5 to about 1.0% of a nonionic surfactant; and centrifuging the mixture; (b) adjusting the pH of the supernatant to 5.5 with 10% acetic acid and centrifuging; (c) eluting the supernatant through a cation-exchange agarose column with 25 mM phosphate buffer; and collecting the desired fractions.
Claim 10
Dependent Claim
29 words
The process of Claim 8, wherein Step (a) is carried out in 10-40 mM calcium chloride solution at a temperature between about 10 and 25°C for about 15-45 minutes.
Claim 11
Dependent Claim
24 words
The process of Claim 8, wherein the elution in Step (b) is carried out with 25-50 mM phosphate buffer containing 0.1 M sodium chloride.
Claim 12
Dependent Claim
31 words
The process of Claim 8, wherein the elution in Step (C) is carried out using a salt gradient of 0.1 M to 1.0 M sodium chloride in 25 mM phosphate buffer.
Claim 13
Dependent Claim
25 words
The process of Claim 9, wherein the desired fractions are further clarified by freezing then thawing up to about 25°; centrifuging, and collecting the supernatant.
References Cited (Backward Citations)
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