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. Author manuscript; available in PMC: 2020 Dec 1.
Published in final edited form as: Nat Immunol. 2019 Oct 21;20(12):1644–1655. doi: 10.1038/s41590-019-0504-0

Fig. 6 |. Lipids blocking CD1d-restricted antigen presentation are increased in thymic lysosomes of 2-week-old Asm−/− mice.

Fig. 6 |

a, Lysosomes were extracted from the thymi of WT and Asm−/− mice using magnetic beads. The figure shows lysosome identity for the samples by western blotting for markers of different cellular compartments in the whole tissue lysate, the magnetic-bead-purified lysosomes, the soluble non-bound material, mitochondria-enriched sediment and the non-magnetic microsomal debris. The results are representative of three independent experiments. The blot images are cropped so all markers can be visualized. b, Electron microscopy images of the purified lysosome fraction. The red arrows indicate lysosomes. The black bar represents 100 nm. The results are representative of three independent experiments. c, Sphingomyelin levels in the magnetic bead fraction were quantified by mass spectrometry. The graphs show the ratios of the indicated sphingomyelin species to the total level of ceramide in 2-week-old WT and Asm−/− mice. d,e, Ratios of the indicated sphingomyelin species to the total level of ceramide in 10-week-old (d) and 18- to 20-week-old (e) WT and Asm−/− mice. In all panels, the mean values are shown with the error bars representing the s.e.m. P values were calculated by a two-sided Student’s t-test (c–e). *P < 0.05, **P < 0.01, ***P < 0.001. UC, ultracentrifuge; CtsD, cathepsin D; Cer, ceramide; SM, sphingomyelin.