Rolling circle amplification-based detection of human topoisomerase I activity on magnetic beads.
Anal Biochem
; 451: 42-4, 2014 Apr 15.
Article
em En
| MEDLINE
| ID: mdl-24525043
ABSTRACT
A high-sensitivity assay has been developed for the detection of human topoisomerase I with single molecule resolution. The method uses magnetic sepharose beads to concentrate rolling circle products, produced by the amplification of DNA molecules circularized by topoisomerase I and detectable with a confocal microscope as single and discrete dots, once reacted with fluorescent probes. Each dot, corresponding to a single cleavage-religation event mediated by the enzyme, can be counted due to its high signal/noise ratio, allowing detection of 0.3pM enzyme and representing a valid method to detect the enzyme activity in highly diluted samples.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA Circular
/
DNA Topoisomerases Tipo I
/
Microscopia Confocal
/
Técnicas de Amplificação de Ácido Nucleico
/
Magnetismo
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Revista:
Anal Biochem
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Itália