Dry reagent for blood coagulation tests

• US 5,059,525 A
• Filed: 05/29/1990
• Issued: 10/22/1991
• Est. Priority Date: 11/19/1984
• Status: Expired due to Fees

• Current Assignee: Boehringer Mannheim GmbH (Roche Holding AG)
• Original Assignee: Boehringer Mannheim GmbH (Roche Holding AG)
• Inventors: Bartl, Knut, Becker, Udo, Lill, Helmut, Wielinger, Hans
• Primary Examiner(s): Kepplinger, Esther L.
• Assistant Examiner(s): Spiegel, Carol A.
• Application Number:
  US07/530,469
• Time in Patent Office: 511 Days
• Field of Search: 435/13, 435/176, 435/177, 435/805, 435/212, 435/214, 435/217, 436/69, 436/166, 436/169, 436/170, 436/810, 530/802, 422/56, 422/57
• US Class Current: 435/13
• CPC Class Codes: C12Q 1/56 involving blood clotting factors, e.g. involving thrombin, thromboplastin, fibrinogen
• CPC Class Codes: G01N 33/525 Multi-layer analytical elements
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick
• CPC Class Codes: Y10S 530/802 Chromogenic or luminescent peptides

Abstract

The present invention provides a dry reagent for blood coagulation tests in which an at least partial course of the coagulation cascade takes place, comprising a carrier material which contains a chromophoric substrate of a protease of the blood coagulation system, at least one factor and/or co-factor of the blood coagulation system and a buffer.

Claims

1. Test strip useful in carrying out a Quick Test, a partial prothromboplastin time test, or in an analysis of one of prothrombin, Factor VII, Factor VIII, Factor IX and Factor X, comprising:

(i) a carrier material containing (ii) a protease which reacts with prothrombin, Factor VII, Factor VIII, Factor IX or Factor X to yield an activated protease;

(iii) a chromophoric protease substrate cleavable by said activated protease in a reaction which does not change the course of the intrinsic or extrinsic coagulation cascade system of blood in the presence of said carrier material, (iv) a buffer, and (v) at least one additional member of the intrinsic or extrinsic coagulation cascade system, wherein said protease reacts with prothrombin, Factor VII, Factor VIII, Factor IX or Factor X in a reaction which is part of the intrinsic or extrinsic coagulation cascade system wherein (ii), (v) are dried on said carrier material.

2. The test strip of claim 1 further comprising a second carrier material comprising an oxidizing agent, and said first carrier material further comprises an aniline or phenol derivative which forms a colored compound with the chromophoric protease substrate after said chromophoric protease substrate has been cleaved by said protease to release a chromophore in the presence of said oxidizing agent.

3. The test strip of claim 2 wherein the aniline or phenol derivative is N-methylanthranilic acid, dimethylanthranilic acid, N-ethyl-N-(3'"'"'-sulphobenzene)-aniline or 2,3- xylenol.

4. The test strip of claim 2, wherein said chromophoric substrate is Tos-Gly-Pro-Arg-p-phenylenediamine, said aniline derivative is N-methylanthranilic acid and said oxidizing agent is potassium ferricyanide.

5. The test strip of claim 2, further comprising a third sample receiving carrier material positioned above or below said first carrier material.

6. The test strip of claim 5, further comprising a fiber fleece arranged between the third and first carrier material and separating said third and first carrier materials from each other.

7. The test strip of claim 5, further comprising a fiber fleece arranged between the first and second carrier material and separating said first and second carrier materials from each other.

8. The test strip of claim 1 useful in carrying out a Quick Test, comprising thromboplastin, a chromophoric substrate for thrombin and Ca²⁺ ions.

9. The test strip of claim 1 useful in analysis of prothrombin, comprising Factor Xa, Factor V, Ca²⁺ ions and a phospholipid.

10. The test strip of claim 1 useful in carrying out a partial thromboplastin time test, comprising a thrombin activator, a contact factor activator, a chromophoric substrate for thrombin, and Ca²⁺ ions.

11. The test strip of claim 10, further comprising ellagic acid and a phospholipid.

12. The test strip of claim 1 useful in analysis of Factor X, comprising at least one factor involved in blood coagulation selected from the group consisting of venom obtained from Russell'"'"'s viper and Factor X activating enzyme derived from Russell'"'"'s viper venom, and further comprising Ca²⁺ ions and a chromophoric substrate for factor Xa .

13. The test strip of claim 1 useful in analysis of Factor VIII, comprising Factor IXa, thrombin, Ca²⁺ ions, a phospholipid and one member of the group consisting of a chromophoric substrate for Factor Xa and a chromophoric substrate for thrombin.

14. The test strip of claim 1 useful in analysis of Factor VII, comprising thromboplastin, Ca²⁺ ions and a chromophoric substrate for Factor Xa.

15. The test strip of claim 1 useful in analysis of factor IX comprising an activated contact factor, Factor IXa, Ca²⁺ ions and one of a chromophoric substrate for Factor IXa, or a combination of phospholipid, Factor VIII, thrombin and a chromophoric substrate for Factor Xa.

16. The test strip of claim 1 wherein said chromphoric substrate is a compound of the formula ##STR3## wherein A is the amino acid arginine or lysine, X is an N-terminal amino acid protective group, Y is a single bond or a chain of 1 to 3 amino acids, NR₁ R₂ is a group in the o- or p-position in which R₁ and R₂, independently of one another, are hydrogen, alkyl containing up to 3 carbon atoms or nitro and R₃ is hydrogen, a carboxyl ester or carboxylamido group, halogen, nitro or an alkyl containing up to 3 carbon atoms.

17. The test strip of claim 16 wherein said chromophoric substrate is a Tos-Gly-Pro-Arg derivative.

18. The test strip of claim 16, further comprising a second carrier material which comprises an oxidizing agent and said first carrier material comprises an aniline or phenol derivative which forms a colored compound with the chromophore of the chromophoric substrate in the presence of said oxidizing agent.

19. The test strip of claim 1, wherein the carrier material is absorbent.