American Society of Hematology

Figure 3.

$Figure 3. Deletion of the intracellular FasL domain abolishes raft localization of the ligand. PNSs from HEK293 cells stably expressing full-length human FasL (A), FasL delta 1-40 (lacking the N-terminal aa 1-40 (B), or FasL delta 1-80 (lacking the N-terminal aa 1-80 that constitutes the complete intracellular domain) (C) were prepared and then solubilized in Brij 98 detergent before they were subjected to sucrose gradient separation, as described in “Materials and methods.” Western blot analysis of the single gradient fractions was performed with anti-FasL, anti-Fyn (raft marker), and anti-Rab5 (non–raft marker) antibodies. Flow cytometry analysis of FasL expression was performed on each cell line to ensure that the deletion mutants FasL delta 1-40 and FasL delta 1-80 were properly expressed at the cell surface. The thin line indicates secondary antibody alone, thick line, Anti–FasL antibody (Nok-1) plus secondary antibody.$

Deletion of the intracellular FasL domain abolishes raft localization of the ligand. PNSs from HEK293 cells stably expressing full-length human FasL (A), FasL delta 1-40 (lacking the N-terminal aa 1-40 (B), or FasL delta 1-80 (lacking the N-terminal aa 1-80 that constitutes the complete intracellular domain) (C) were prepared and then solubilized in Brij 98 detergent before they were subjected to sucrose gradient separation, as described in “Materials and methods.” Western blot analysis of the single gradient fractions was performed with anti-FasL, anti-Fyn (raft marker), and anti-Rab5 (non–raft marker) antibodies. Flow cytometry analysis of FasL expression was performed on each cell line to ensure that the deletion mutants FasL delta 1-40 and FasL delta 1-80 were properly expressed at the cell surface. The thin line indicates secondary antibody alone, thick line, Anti–FasL antibody (Nok-1) plus secondary antibody.

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