Patent US5618684A successfully refreshed from Google Patents.
US5618684A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Method of determination of calcium
Assignee: Oriental Yeast Co Ltd
•
Filing: 1995-04-20
•
Granted: 1997-04-08
•
Expires: 2015-04-20
Abstract
Abstract Calcium in a sample is brought into contact with a transglutaminase capable of being activated with calcium as an activating factor and the transglutaminase activity, which varies depending upon the calcium amount in the sample, is measured to thereby determine the calcium amount in the sample. By the method of the invention, accurate determination of calcium in various samples such as body fluids is possible without removal of proteins from them.
Inventors
- 👤 Masatsugu Nonobe
- 👤 Hozumi Nishida
- 👤 Tsuyoshi Fujita
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-30 16:47
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/48
Claims Analysis
Total Claims: 18
Claim 1
Independent Claim
115 words
A method of determining the amount of calcium in blood serum, comprising the steps of: (1) providing a sample of blood serum; (2) bringing blood serum calcium in said sample of blood serum into contact with a transglutaminase capable of being activated with calcium to provide an activated transglutaminase; (3) allowing the activated transglutaminase to act on a substrate which is a mixture of a donor and an acceptor to generate NH 3 ; and (4) measuring the activity of transglutaminase by detecting the amount of NH 3 generated; and (5) determining the amount of calcium in the sample of blood serum by correlating the amount of NH 3 generated with an amount of calcium.
Claim 2
Independent Claim
115 words
A method of determining the amount of calcium in blood serum, comprising the steps of: (1) providing a sample of blood serum; (2) bringing blood serum calcium in said sample of blood serum into contact with a transglutaminase capable of being activated with calcium to provide an activated transglutaminase; (3) allowing the activated transglutaminase to act on a substrate which is a mixture of a donor and an acceptor to generate NH 3 ; and (4) measuring the activity of transglutaminase by detecting the amount of NH 3 generated; and (5) determining the amount of calcium in the sample of blood serum by correlating the amount of NH 3 generated with an amount of calcium.
Claim 3
Independent Claim
119 words
A method of determining the amount of calcium in blood serum, comprising the steps of: (1) providing a sample of blood serum; (2) bringing blood serum calcium in said sample of blood serum into contact with a transglutaminase capable of being activated with calcium to provide an activated transglutaminase; (3) allowing the activated transglutaminase to act on a substrate which is a mixture of a donor and an acceptor, wherein said acceptor is hydroxylamine, to generate hydroxamic acid; and (4) measuring the activity of transglutaminase by detecting the amount of hydroxamic acid generated; and (5) determining the amount of calcium in the sample of blood serum by correlating the amount of hydroxamic acid generated with an amount of calcium.
Claim 4
Independent Claim
119 words
A method of determining the amount of calcium in blood serum, comprising the steps of: (1) providing a sample of blood serum; (2) bringing blood serum calcium in said sample of blood serum into contact with a transglutaminase capable of being activated with calcium to provide an activated transglutaminase; (3) allowing the activated transglutaminase to act on a substrate which is a mixture of a donor and an acceptor, wherein said acceptor is hydroxylamine, to generate hydroxamic acid; and (4) measuring the activity of transglutaminase by detecting the amount of hydroxamic acid generated; and (5) determining the amount of calcium in the sample of blood serum by correlating the amount of hydroxamic acid generated with an amount of calcium.
Claim 5
Dependent Claim
79 words
A method according to claim 1, wherein said donor is a compound represented by the formula, X-L-Gln-Y, wherein X is an amino acid, peptide or protective group at the N-terminal end of L-Gln, and Y is an amino acid, peptide or hydrogen atom at the C-terminal end of L-Gln, and said acceptor is a compound represented by the formula, R-NH.sub.2, wherein R is (CH 2 ) n CH 3 where n=2-5, OH or lysine residue (R-NH 2 =lysine ).
Claim 6
Dependent Claim
49 words
A method according to claim 2, wherein said donor is a compound represented by the formula, X-L-Gln-Y, wherein X is an amino acid, peptide or protective group at the N-terminal end of L-Gln, and Y is an amino acid, peptide or hydrogen atom at the C-terminal end of L-Gln.
Claim 7
Dependent Claim
37 words
A method according to claim 1, wherein said donor is selected from the group consisting of benzyloxycarbonyl-L-Gln-Gly, benzyloxycarbonyl-L-Gln, t-butyloxycarbonyl-L-Gln, and 9-fluorenylmethoxycarbonyl-L-Gln, and said acceptor is selected from the group consisting of n-propylamine, n-butylamine, n-amylamine, n-hexylamine, and lysine.
Claim 8
Dependent Claim
21 words
A method according to claim 2, wherein said donor is selected from the group consisting of benzyloxycarbonyl-L-Gln-Gly, benzyloxycarbonyl-L-Gln, t-butyloxycarbonyl-L-Gln, and 9-fluorenylmethoxycarbonyl-L-Gln.
Claim 9
Dependent Claim
37 words
A method according to claim 3, wherein said donor is selected from the group consisting of benzyloxycarbonyl-L-Gln-Gly, benzyloxycarbonyl-L-Gln, t-butyloxycarbonyl-L-Gln, and 9-fluorenylmethoxycarbonyl-L-Gln, and the acceptor is selected from the group consisting of n-propylamine, n-butylamine, n-amylamine, n-hexylamine, and lysine.
Claim 10
Dependent Claim
21 words
A method according to claim 4, wherein said donor is selected from the group consisting of benzyloxycarbonyl-L-Gln-Gly, benzyloxycarbonyl-L-Gln, t-butyloxycarbonyl-L-Gln, and 9-fluorenylmethoxycarbonyl-L-Gln.
Claim 11
Dependent Claim
27 words
A method according to claim 1, further comprising the step of pre-incubating said transglutaminase and said substrate separately at about 37° C. prior to activating said transglutaminase.
Claim 12
Dependent Claim
23 words
A method according to claim 1, wherein step (2) is carried out at a temperature of about 37° C. for about two minutes.
Claim 13
Dependent Claim
22 words
A method according to claim 9, wherein said transglutaminase is used in said pre-incubation step in an amount of about 10 U/ml.
Claim 14
Dependent Claim
27 words
A method according to claim 2, further comprising the step of pre-incubating said transglutaminase and said substrate separately at about 37° C. prior to activating said transglutaminase.
Claim 15
Dependent Claim
22 words
A method according to claim 2, wherein step (2) is carried out at a temperature of about 37° C. for two minutes.
Claim 16
Dependent Claim
22 words
A method according to claim 12, wherein said transglutaminase is used in said pre-incubation step in an amount of about 10 U/ml.
Claim 17
Dependent Claim
47 words
The method according to claim 1, wherein the amount of calcium in the sample of blood serum is determined by calculating from a standard curve the amount of calcium in the sample that corresponds to the amount of NH 3 detected as a measure of transglutaminase activity.
Claim 18
Dependent Claim
47 words
The method according to claim 2, wherein the amount of calcium in the sample of blood serum is determined by calculating from a standard curve the amount of calcium in the sample that corresponds to the amount of hydroxamic acid detected as a measure of transglutaminase activity.
References Cited (Backward Citations)
JPS5931698A
Cited
JPS61247963A
Cited
JPS626699A
Cited
JPS626700A
Cited
JPS6236199A
Cited
JPS642598A
Cited
US5015588A
Cited
US5049506A
Cited
JPH01231896A
Cited
US5204240A
Cited
US5204447A
Cited
JPH02255098A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 2000-04-07 | 2000-10-07 | 2001-04-08 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2004-04-07 | 2004-10-07 | 2005-04-08 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2008-04-07 | 2008-10-07 | 2009-04-08 | $7700.00 | Window Closed / Lapsed |