Analysis of internucleosomal DNA fragmentation in apoptotic thymocytes by dynamic sieving capillary electrophoresis.
The analysis, by slab gel electrophoresis, of internucleosomal DNA cleavage or laddering, characteristic of apoptosis in many cell systems, is labour intensive, difficult to automate and at best only semi-quantitative. In this report we show that CE, using dilute solutions of hydroxyethylcellulose as a replaceable sieving matrix, can be applied to the relatively rapid analysis of DNA laddering in whole digests of apoptotic rat thymocytes. Also, using the sensitivity of laser-induced fluorescence detection and the highly sensitive nuclei acid stain YO-PRO-1, the CE method reported here can use 1000-2000 fold fewer cells than needed for traditional slab gel methods.
Evans
M D
MD
Division of Chemical Pathology, University of Leicester, UK.
Wolfe
J T
JT
Perrett
D
D
Lunec
J
J
Herbert
K E
KE
D000818:Animals; D017209:Apoptosis; D002197:Capillary Action; D002458:Cell Fractionation; D004247:DNA / Q000032:analysis*; D004586:Electrophoresis / Q000379:methods*; D019286:Endopeptidase K; D008297:Male; D009707:Nucleosomes / Q000737:chemistry*; D051381:Rats; D011916:Rats, Inbred F344; D012260:Ribonucleases / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D013950:Thymus Gland / Q000737:chemistry* / Q000648:ultrastructure
D009707:Nucleosomes; D004247:DNA; D012260:Ribonucleases; D012697:Serine Endopeptidases; D019286:Endopeptidase K