Fig. 4

Fluorescence confocal imaging visualizes the effects of the in situ isolation of nuclear proteins from adherent nuclei on the cytoplasmic and intranuclear contents of HUVEC cells. a Dynamic observation of the morphological and fluorescence changes of cells after successive Triton X-100 (0.1%) and SDS (0.1%) treatments as indicated by the black arrows. Upper panels: DIC images (the imaging focus is on cell morphology); lower panels: images merged from DIC and fluorescence images (the imaging focus is on fluorescence). The cytoplasmic contents were stained with a red fluorescent dye (SQ-2) and the intranuclear chromosomes were stained with Hoechst33342 (blue). b Depleting effect of 0.1% Triton X-100 for 10 min on tubulin cytoskeleton. Left panels: images merged from DIC and fluorescence images (tubulin cytoskeleton in green and the nuclei in blue); right panels: DIC images. c, d No effects of 0.1% Triton X-100 on intranuclear contents including chromosomes and proteins (e.g., pP65 in c and Hif-1α in d as representatives). The HUVEC cells have been activated by 1 μg/mL LPS prior to Triton treatment. The chromosomes were stained with DAPI (blue), pP65 was stained with anti-pP65 antibody and AlexaFluor647-conjugated anti-IgG antibody (red), and Hif-1α was stained with anti-Hif-1α mAb and FITC-conjugated goat anti-mouse IgG. Scale bars: 20 μm (a–d). Additional file 2: Video S1 also shows the time-lapse observation of the effects of successive Triton X-100 and SDS treatments on the mitochondria and nuclei of cells fluorescently stained with Mitotracker (red) and Hoechst33342 (blue)