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. 1997 Dec 15;139(6):1433–1446. doi: 10.1083/jcb.139.6.1433

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(a) Sedimentation profiles of endosomal and lysosomal organelles after isopycnic centrifugation is sucrose. Mel JuSo cells were allowed to internalize HRP for 5 min followed by a 15-min chase. Cells were homogenized and a post-nuclear supernatant was loaded on a sucrose gradient (0.8–1.2 M sucrose in homogenization buffer). After centrifugation and fractionation of the gradients (left top, right bottom), each fraction was analyzed for the activity of β-hexosaminidase (β-hex, top) and HRP (middle). (Bottom) Total protein content in the different fractions as analyzed by Bradford. Shown are mean (± SD) from three separate experiments. (b) Steady state distribution of MHC class II, Ii and HLA-DM molecules in subcellular organelles. Proteins from each fraction after sucrose density gradient centrifugation (left top, right bottom) were allowed to bind to nitrocellulose after which the amount of MHC class II (top), Ii (middle) or HLA-DM (bottom) was analyzed by immunostaining, using specific antibodies. Shown are mean (± SD) obtained from three separate gradients.