An ultrafiltration assay for lysyl oxidase
- Univ. of Edinburgh Medical School (England)
A modification of the original microdistillation assay for lysyl oxidase is described in which Amicon C-10 microconcentrators are used to separate, by ultrafiltration, the 3H-labeled products released from a (4,5-3H)-lysine-labeled elastin substrate. Enzyme activity is determined by scintillation counting of the ultrafiltrate, after subtraction of radioactivity released in the presence of beta-aminopropionitrile, a specific inhibitor of the enzyme. Conditions are described which optimize both the sensitivity and the efficient use of substrate. The assay shows linear inhibition of activity in up to 1 M urea; hence, as the enzyme is normally diluted in the assay, samples in 6 M urea can be assayed directly, without prior dialysis, and corrected for partial inhibition. Comparable results are obtained when enzyme activity is assayed by ultrafiltration or microdistillation. The assay is simple and convenient and, by using disposable containers throughout, it eliminates the need for time-consuming decontamination of radioactive glassware.
- OSTI ID:
- 6749547
- Journal Information:
- Analytical Biochemistry; (USA), Journal Name: Analytical Biochemistry; (USA) Vol. 185:2; ISSN ANBCA; ISSN 0003-2697
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
AMIDES
AMINO ACIDS
ANIMALS
BIRDS
CARBONIC ACID DERIVATIVES
CARBOXYLIC ACIDS
CHICKENS
COUNTING TECHNIQUES
DOMESTIC ANIMALS
DOSE-RESPONSE RELATIONSHIPS
EMBRYOS
ENZYME INHIBITORS
ENZYMES
FILTRATION
FOWL
HYDROGEN COMPOUNDS
MAMMALS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXIDOREDUCTASES
PROTEINS
RADIOASSAY
SCINTILLATION COUNTING
SEPARATION PROCESSES
SWINE
TRITIUM COMPOUNDS
UREA
VERTEBRATES