Detection of DNA Fragmentation in Apoptotic Cells by TUNEL.
Cold Spring Harb Protoc
; 2016(10)2016 10 03.
Article
em En
| MEDLINE
| ID: mdl-27698233
ABSTRACT
Degradation of DNA into oligonucleosomal-sized fragments is a unique event in apoptosis that is orchestrated by caspase-activated DNase. Traditionally, this event is observed by resolving cellular DNA by gel electrophoresis, which results in a characteristic "ladder" pattern. However, this technique is time-consuming and cannot be used to quantitate the number of apoptotic cells in a sample. Terminal dUTP nick-end labeling (TUNEL) of fragmented DNA allows researchers to identify DNA fragmentation at the single-cell level. This method involves the specific addition of fluorescently labeled UTP to the 3'-end of the DNA fragments by terminal deoxynucleotidyl transferase. The TUNEL assay is both fast and sensitive. Here, we describe a protocol in which cells are treated with TUNEL reagent and counterstained with Hoechst 33342. In contrast to TUNEL, which only stains apoptotic cells, Hoechst 33342 stains the DNA of all cells.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Técnicas Citológicas
/
Apoptose
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Marcação In Situ das Extremidades Cortadas
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Fragmentação do DNA
Tipo de estudo:
Diagnostic_studies
/
Prognostic_studies
Idioma:
En
Revista:
Cold Spring Harb Protoc
Ano de publicação:
2016
Tipo de documento:
Article
País de afiliação:
Austrália