Fig. 2. Nuclear localization of PrP^Sc in prion-infected N2a cells by immunofluorescence. N2a and ScN2a cells were fixed, permeabilized, treated with 3M guanidine thiocyanate and processed for immunofluorescence labelling with antibodies against PrP (SAF61 antibody), followed by rhodamine-conjugated anti-mouse secondary antibodies. Cells were viewed with blue excitation/emission settings to detect Hoechst staining of the nuclei (e,f,g,h) and with red excitation/emission settings to detect PrP (a,b,c,d). (A) Non-infected N2a cells (a,e) or prion-infected cells (b-d,f-h) were observed by indirect immunofluorescence on a Leica microscope. (Ab) White arrows indicate cells that have a nuclear accumulation of PrP^Sc. (B) Staining of ScN2a cells was analyzed by confocal laser-scanning microscopy; arrow in panels i-k: localization of PrP in the nucleus is visible in this optical section of the cells (g-i). Bars, 8 µm.