Biotinylated deoxyadenosine triphosphate (dATP) (Bio7dATP) and 3H deoxythymidine triphosphate (dTTP) labeled adenovirus DNA were hybridized in situ to thin sections of Lowicryl K4Membedded and wholemount extracted HeLa cells infected with adenovirus. The biotinylated probe was detected by exposing the extracted cells or sections to antibodies against biotin followed by colloidal goldconjugated secondary antibodies and then criticalpoint dried while 3HdTTP labeled probe by electron microscopic autoradiography. On Lowicryl K4M sections, gold particles and silver grains were mainly restricted in the nucleus. Furthermore, wholemount results suggested that replicating adenovirus DNA is localized on the nuclear matrix of its host cell. In this paper, the described nonradioactive procedures for hybrid detection offered several advantages: (a) rapid signal detection; (ob) superior morphological preservation and spatial resolution; (c) precise localization; and (d) on Lowicryl K4M sections, signal to noise equivalent to radiolabeling. Copyright1992 WileyLiss, Inc.
- DNA replication
- Nick translation
- Nuclear matrix
ASJC Scopus subject areas
- Medical Laboratory Technology