Subnuclear immunofluorescence.
Despite their relatively small size and the fact that they are naturally non-adherent, it is possible to obtain good immunofluorescence staining of subnuclear structures in DT40. This, combined with their genetic tractability, provides a powerful combination for the study of DNA replication and repair. Here we provide a general protocol for immunofluorescence of molecules such as RAD51 and phosphorylated histone H2Ax. We also present a modification to this protocol that allows visualisation of chromatin bound PCNA and hence sites of DNA synthesis.
Szüts
David
D
M.R.C. Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH.
Sale
Julian E
JE
D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D005455:Fluorescent Antibody Technique / Q000379:methods*; D010766:Phosphorylation