Determination of total creatine kinase or an isoenzyme with a multilayer analytical element

• US 4,713,327 A
• Filed: 05/01/1985
• Issued: 12/15/1987
• Est. Priority Date: 05/01/1985
• Status: Expired due to Fees

• Current Assignee: Clinical Diagnostic Systems Inc.
• Original Assignee: Eastman Kodak Company
• Inventors: Findlay, John B., Wu, Annie L., Norton, Gary E.
• Primary Examiner(s): Naff, David M.
• Application Number:
  US06/729,331
• Time in Patent Office: 958 Days
• Field of Search: 435/17, 435/291, 422/56, 422/57, 436/528, 436/535, 436/169
• US Class Current: 435/17
• CPC Class Codes: C12Q 1/50 involving creatine phosphokinase
• CPC Class Codes: G01N 33/526 the element being adapted for a specific analyte
• CPC Class Codes: G01N 33/573 for enzymes or isoenzymes

Abstract

A multilayer analytical element for the determination of creatine kinase comprises a support having thereon, in order and in fluid contact, a registration layer and an isotropically porous spreading layer. The registration layer contains a binder material at a coverage of at least about 8 g/m², which coverage improves the stability and keeping properties of the element in high humidity environments. This element is useful in an analytical method for determining total creatine kinase or any one of its isoenzymes, e.g. creatine kinase-MB.

Claims

1. A multilayer analytical element for the determination of total creatine kinase or a creatine kinase isoenyzme, said element comprising three or more layers and containing in one or more of said layers, a substrate for creatine kinase, said element also comprising a support having thereon,

a registration layer containing a binder material at a coverage of at least about 8 g/m² and an indicator composition which provides a detectable change in response to the reaction of said substrate with said creatine kinase or said isoenzyme,

a reagent layer, and

an isotropically porous spreading layer containing an activator for creatine kinase, said layers being in order from said support and in fluid contact.

2. The element of claim 1 wherein said indicator composition provides an optical density change in response to adenosine-5'"'"'-triphosphate or its reaction product.

3. The element of claim 1 wherein said binder material is present at a coverage of at least about 10 g/m².

4. The element of claim 1 wherein said binder material is gelatin.

5. The element of claim 1 wherein said reagent layer contains creatine phosphate.

6. The element of claim 1 wherein said reagent layer comprises adenosine-5'"'"'-monophosphate, adenosine-5'"'"'-diphosphate and P¹, P⁵ -di(adenosine-5'"'"')pentaphosphate.

7. The element of claim 1 wherein said porous spreading layer contains one or more antibodies for an isoenzyme of creatine kinase.

8. A multilayer analytical element for the determination of creatine kinase-MB, said element comprising a nonporous support having thereon, in order from said support and in fluid contact,

a registration layer containing a binder material at a coverage of at least about 10 g/m², and an indicator composition which provides an optical density change in response to adenosine-5'"'"'-triphosphate or its reaction product,

a reagent layer containing creatine phosphate, adenosine-5'"'"'-diphosphate, adenosine-5'"'"'-monophosphate and P¹,P⁵ -di(adenosine-5'"'"')pentaphosphate, and

an isotropically porous spreading layer containing an activator for creatine kinase and one or more antibodies for creatine kinase-MM.

9. The element of claim 8 wherein said registration layer binding material is gelatin.

10. The element of claim 8 wherein said adenosine-5'"'"'-diphosphate is present in an amount of from about 0.05 to about 0.16 g/m², said adenosine-5'"'"'-monophosphate is present in a molar ratio to adenosine-5'"'"'-diphosphate of at least about 10:

1, and P¹,P⁵ -di(adenosine-5'"'"')pentaphosphate is present in an amount of at least about 0.03 g/m².

11. The element of claim 8 wherein said registration layer contains α

-glycerophosphate oxidase, said reagent layer contains glycerol and glycerol kinase, and said indicator composition comprises a substance having peroxidative activity and a leuco dye.

16. A method for the determination of creatine kinase-MB in an aqueous liquid, said method comprising the steps of:

(A) contacting a sample of said liquid with the analytical element of claim 8, and

(B) determining the resulting optical density change.

17. The method of claim 16 wherein said element comprises adensoine-5'"'"'-diphosphate present in an amount of from about 0.05 to about 0.16 g/m², adensoine-5'"'"'-monophosphate present in a molar ratio to adenosine-5'"'"'-diphosphate of at least about 10:

1, and P¹, P⁵ -di(adenosine-5'"'"') pentaphosphate present in an amount of at least about 0.03 g/m².

18. The method of claim 16 wherein said element has α

-glycerophosphate in the registration layer, glycerol and glycerol kinase in the reagent layer, and the indicator composition comprises a substance having peroxidative activity and a leuco dye.

19. The method of claim 16 wherein said element contains gelatin as said binder material in said registration layer.

12. A method for the determination of total creatine kinase or a creatine kinase isoenzyme in an aqueous liquid, said method comprising the steps of:

(A) contacting a sample of said liquid with an analytical element comprising three of more layers and containing in one or more of said layers, a substrate for creatine kinase, said element also comprising a support having thereon,

a registration layer containing a binder material at a coverage of at least about 8 g/m² and an indicator composition which provides a detectable change in response to the reaction of said substrate with said creatine kinase or said isoenzyme,

a reagent layer, and

an isotropically porous spreading layer containing an activator for creatine kinase, said layers being in order from said support and in fluid contact, and

(B) determining the resulting detectable change.

13. The method of claim 12 for the determination of creatine kinase-MB wherein said contact is carried out in the presence of one or more antibodies for creatine kinase-MM.

14. The method of claim 12 wherein said detectable change is an optical density change.

15. The method of claim 14 wherein said optical density change is a colorimetric change.