Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving transferase
Patent
1995-08-18
1998-11-10
Kunz, Gary L.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving transferase
435 26, 435189, 435194, 536 2622, 536 2624, 562553, C12Q 150, C12Q 132, C12N 904, C12N 912
Patent
active
058342273
DESCRIPTION:
BRIEF SUMMARY
This case is a 371 of PCT/JP94/02136 filed Dec. 19, 1994.
TECHNICAL FIELD
The present invention relates to a reagent for assaying creatine kinase (hereinafter referred to as CK) in a sample, more particularly, an improved liquid reagent for assaying CK, which is stable over a long period.
BACKGROUND ART
CK is distributed in skeletal, cardiac and smooth muscles and a brain. CK is an important enzyme taking part in energy metabolism, and is released into blood from tissues when a muscle or heart is suffered from disease. It is important to assay CK activity as indices for various diseases.
CK catalyzes following reaction: phosphate.fwdarw.ATP+creatine (reverse reaction) wherein ATP is adenosine 5'-triphosphate, and ADP is adenosine 5'-diphosphate.
In the assay of CK, the products of the forward and reverse reactions are used. According to the reaction product to be determined, the assay is broadly classified into the following four methods: reduced nicotinamide adenine dinucleotide (NADH) is measured, using pyruvate kinase and lactate dehydrogenase as conjugated enzymes. phosphorus produced by hydrolyzing creatine phosphate is measured. reduced nicotinamide adenine dinucleotide phosphate (hereinafter referred to as NADPH) is measured, using mainly hexokinase (hereinafter referred to as HK) or glucokinase (hereinafter referred to as GlcK) and glucose-6-phosphate dehydrogenase (hereinafter referred to as G6PDH) as conjugated enzymes. from creatine produced from the CK enzyme reaction is measured by means of Jaffe's method or the like.
Of the above methods (1) to (4), the method (3) for determining ATP has been widely used, and recommended as a standard method for determining CK activity by Japanese Society of Clinical Chemistry (JSCC).
The reactions for determining CK activity in HK-G6PDH method which is a typical example of the method (3) for determining ATP are as follows: ##STR1## wherein NADP.sup.+ is oxidized nicotinamide adenine dinucleotide phosphate.
The HK-G6PDH method is very excellent for determining CK activity. However, very unstable enzymes and substrates were used, and thus, a long term storage was difficult. Therefore, the enzymes, substrates and the like are supplied in the form of lyophilized products. Further, stabilizing conditions for the reagents widely vary with their combinations. Therefore, it was difficult to maintain stability of liquid reagent for a long period of time by conventional combinations.
With the spread of an automatic analyzer, the reagent composition used in the method for determining CK activity has been converted from a one-component form to a two-component form. According to the conventional combination of the two-component form, a reagent for initiating the reaction (second reagent) which is a substrate for determining CK generally contains creatine phosphate, and a first reagent contains all the remaining components. However, the storage stability as a liquid reagent was very poor. Further, some second reagents contained HK and/or G6PDH, there was a problem in the stabilities of the enzymes. Furthermore,. it was not possible to obtain sufficient stability by other combinations.
Recently, it is desired to improve the workability for users, by providing the reagents in a liquid form from a supplier, instead of preparing the liquid reagent when used.
The stability of the enzymes per se has been improved by removing the interfering contaminant enzymes by sophisticated purification of the enzymes or by using heat resistant enzymes, for example, glucokinase (hereinafter referred to as GlcK) derived from Bacillus stearothermophilus, HK obtained from recombinant yeast or chemically modified enzymes thereof. However, it was difficult to obtain a sufficient storage stability for liquid reagents by such enzymes. Particularly, the inventors of the present invention found that, in the reagent used in HK-G6PDH method, the enzymes and substrates which are stable alone respectively, become unstable when contained with each other in a same liquid. Namely, the inventors found
REFERENCES:
patent: 5587296 (1996-12-01), Tsubota
Chemical Abstract 108: 71027, Kondo et al., J. Clin. Biochem. Nutr. 3(1):17-25, 1987.
Shimada Reiko
Tsubota Hiroyuki
Iatron Laboratories Inc.
Kunz Gary L.
LandOfFree
Kit for assaying creatine kinase does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Kit for assaying creatine kinase, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Kit for assaying creatine kinase will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-1515410