Chapter title |
Zymographic Determination of Intrinsic Specific Activity of Oxidases in the Presence of Interfering Proteins
|
---|---|
Chapter number | 24 |
Book title |
Protein Gel Detection and Imaging
|
Published in |
Methods in molecular biology, August 2018
|
DOI | 10.1007/978-1-4939-8745-0_24 |
Pubmed ID | |
Book ISBNs |
978-1-4939-8744-3, 978-1-4939-8745-0
|
Authors |
Tien Canh Le, Mircea Alexandru Mateescu, Samaneh Ahmadifar, Lucia Marcocci, Paola Pietrangeli, Le, Tien Canh, Mateescu, Mircea Alexandru, Ahmadifar, Samaneh, Marcocci, Lucia, Pietrangeli, Paola |
Abstract |
Zymography on peroxidase-entrapped gels enables the determination of the intrinsic specific activity (ISA) of H2O2-producing oxidases in the presence of interfering enzymes including catalase (decomposing hydrogen peroxide). To reveal the searched oxidase, the zymography gel is incubated first in the developing solution containing the appropriate substrate of the oxidase and an oxidizable chromogen such as o-phenylenediamine (o-PDA) peroxidase substrate. The zymographic gel, after image analysis, can be restained by Coomassie Blue for molecular weights, for the whole electrophoretic pattern and for the protein concentration, thus allowing for the determination of oxidase ISA. |
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