Fig. 2. Myoblast differentiation is accompanied by redistribution of lamins. C2C12 myoblasts were grown on glass coverslips and induced to differentiate with 2% horse serum. 0, 48-72 and 96-120 h after transfer to differentiation medium, cells were fixed and co-stained for either lamin A/C (a) or lamin B1(b) and myogenin (N.B. myoblasts do not differentiate synchronously and therefore the intensity of nuclear staining with anti-myogenin antibodies varies between cells within a single time point). z-axis sections of 48-hour myoblast cultures stained for lamin C were collected at 0.35 µm intervals and projected (c) (short arrows indicate a postmitotic myoblast, asterisks indicate mitotic myoblasts). Alternatively, z-axis series were obtained using a BioRad Lasersharp 2000 confocal microscope with the laser power adjusted to eliminate saturation of the image. Central sections were converted to PICT images without adjustment and surface plotting was performed using LaserPix software. (d) Original midsections stained with anti-lamin-A/C antibodies. Anti-myogenin staining was used to identify differentiated cells (arrows) but is not shown here. Surface plots of the same field are shown in (e). Red coloration shows the areas of most intense fluorescence (maximum output of 215 on a scale of 0-255), whereas blue shows the areas of least intense fluorescence (minimum output 0 on a scale of 0-255). 48-hour cultures were co-stained with anti-lamin-C and anti-lamin-B₂ antibodies. Arrowheads indicate cells showing diffuse anti-lamin-C staining and weak anti-lamin-B₂ staining (f). Images were collected on a BioRad Lasersharp 2000 confocal microscope. Bars, 10 µm.