Patent US2467950A successfully refreshed from Google Patents.
Patent EP0103784A2 successfully refreshed from Google Patents.
EP0103784A2
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Catalyzed colorimetric and fluorometric substrates for peroxidase enzyme determinations
Assignee: ENZYME TECHNOLOGY Co
•
Filing: 1983-08-27
•
Granted: —
•
Expires: 2003-08-27
Abstract
Abstract lmproved, catalyzed substrates for use in developing characteristic colors of florescence in the presence of peroxidase enzymes (e.g., horseradish peroxidase) are disclosed which include as a rate accelerator a substituted phenol such as a p-halogenated phenol. The complete substrate typically includes a peroxide type oxidizing agent (e.g., hydrogen peroxide), a chromogenic or fluorogenic compound (e.g., ABTS), a buffer and the accelerator compound. Advantageously, the accelerator should provide at least about 50 percent rate enhancement for the substrate, as compared with an otherwise identical, accelerator-free substrate reacted under the same conditions; however, the most preferred accelerator, p-iodophenol, gives enhancement on the order to 1,000 percent. The substrates are particularly useful in so-called ELISA determinations which involve an enzyme-linked moiety, and permit detection at very low concentration levels unobtainable with conventional colorimetric substrates.
Inventors
- 👤 Robert L. Stout
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-24 04:27
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/26; C12Q1/28; C12Q2326/00; C12Q2326/30; Y; Y10; Y10S; Y10S435/00; Y10S435/805; Y10S436/00; Y10S436/826; Y10S436/904; Y10T; Y10T436/00; Y10T436/20; Y10T436/206664
Claims Analysis
Total Claims: 11
Claim 1
Independent Claim
88 words
In a substrate for developing a characteristic color or florescence in the presence of a peroxidase enzyme, said substrate including a peroxide oxidizing agent, and a compound capable of reacting and giving off color or florescence in the presence of said enzyme and said oxidizing agent, the improvement which comprises an accelerator admixed with said substrate, taken from the group consisting of substituted phenol compounds, and providing at least about 50% acceleration as compared with an otherwise identical substrate free of said accelerator and reacted under identical conditions.
Claim 2
Dependent Claim
88 words
In a substrate for developing a characteristic color or florescence in the presence of a peroxidase enzyme, said substrate including a peroxide oxidizing agent, and a compound capable of reacting and giving off color or florescence in the presence of said enzyme and said oxidizing agent, the improvement which comprises an accelerator admixed with said substrate, taken from the group consisting of substituted phenol compounds, and providing at least about 50% acceleration as compared with an otherwise identical substrate free of said accelerator and reacted under identical conditions.
Claim 3
Dependent Claim
87 words
The substrate as set forth in Claim 1, said accelerator being taken from the group consisting of vanillin and compounds of the formula wherein R 1 is a halogen, a halogen-substituted. phenoxy compound, a hydroxyphenone or a carboxylic acid or alkyl ester thereof; R 2 is hydrogen, hydroxy, a halogen, substituted and unsubstituted alkyl groups having from 1-6 carbon atoms, inclusive, an amino group, or a carboxylic acid or alkyl ester thereof; and R 3 , R 4 and R 5 are respectively a halogen or hydrogen.
Claim 4
Dependent Claim
41 words
The substrate as set forth in Claim 2, said accelerator being selected from the group consisting of 2,5-dichlorophenol, 2,6-difluoroaniline, 3',3,5 triiodo-L-thyronine, m-bromophenol, o-chlorophenol, 4,4' dihydroxy-benzophenone, 4-fluorophenol, 3,5 diiodo-D-thyronine, 2-3 dichlorophenol, 4-chloro-2-methylphenol, 3,4-dichlorophenol, 3-iodo-L-tyrosine, 2,4-dibromophenol, p-chlorophenol, p-bromophenol, 2,4-dichlorophenol, vanillin, 4-chloro-3-methylphenol and 4-iodophenol.
Claim 5
Dependent Claim
21 words
The substrate as set forth in Claim 3, said accelerator being selected from the group consisting of vanillin, 4-chloro-3-methylphenol and 4-iodophenol.
Claim 6
Dependent Claim
14 words
The substrate as set forth in Claim 1, said oxidizing agent being hydrogen peroxide.
Claim 7
Dependent Claim
18 words
The substrate as set forth in Claim 1, said substrate comprising: 2,2'azino'di-(3-ethyl-benzthozolene-6-sulfonic acid; hydrogen peroxide; 4-iodophenol; and buffer.
Claim 8
Dependent Claim
27 words
The substrate as set forth in Claim 1, said compound being selected from the group consisting of 2,2'-azino-di-(3-ethyl-benzthiozolane-6-sulfonic leid), phenol red, o-phenylenediamine, pyrogallol, 4-aminoantipyrine and bromopyrogallol red.
Claim 9
Independent Claim
85 words
A method of determining the presence of a peroxidase enzyme in a sample comprising the steps of contacting said sample with a substrate including a peroxide oxidizing agent, a compound capable of reacting and giving off color or florescence in the presence of said agent and said enzyme, and an accelerator taken from the group consisting of substituted phenol compounds, said accelerator providing at least about 50% rate acceleration as compared with an otherwise identical substrate free of said accelerator and reacted under identical conditions.
Claim 10
Independent Claim
85 words
A method of determining the presence of a peroxidase enzyme in a sample comprising the steps of contacting said sample with a substrate including a peroxide oxidizing agent, a compound capable of reacting and giving off color or florescence in the presence of said agent and said enzyme, and an accelerator taken from the group consisting of substituted phenol compounds, said accelerator providing at least about 50% rate acceleration as compared with an otherwise identical substrate free of said accelerator and reacted under identical conditions.
Claim 11
Dependent Claim
38 words
The method of Claim 7, said accelerator being selected from the group consisting of 2,5-dichlorophenol, 2,6-difluoroaniline, 3',3,5 triiodo-L-thyronine, m-bromophenol, o-chlorophenol, 4,4' dihydroxy-benzophenone, 4-fluorophenol, 3,5. diiodo-D-thyronine, 2-3 dichlorophenol, 4-chloro-2-methylphenol, 3,4-dichlorophenol, 3-iodo-L-tyrosine, 2,4-dibromophenol, p-chlorophenol, p-bromophenol, 2,4-dichlorophenol, vanillin, 4-chloro-3-methylphenol and 4-iodophenol.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
Maintenance schedule is calculated for granted US utility patents.