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10.1038/ng.2396 108 PisoschiA

High prevalence of fulminant hepatic failure among patients with mutant alleles for truncation of ATP7B in Wilsons disease

For fluorescence microscopy studies, HL-60 cells and HepG2 cells (seeding densities 1 10 4 cells/well and 8 10 4 cells/well, respectively) were placed in serum-free medium to facilitate their adhesion to coverslips and cultured on lysine-coated 8-well chamber slides (IBIDI) following the procedure detailed by 2.8 Cell surface binding, endocytosis, and downregulation of TfRs For surface binding and endocytosis of AF-labeled proteins, the cells were rinsed twice with ice-cold HEPES-buffered, serum-free media, pH 7.4, and incubated for 1 h on ice with either 500 nM hemeAF546HPX, hemeAF546HPX, or AF488 holo-human Tf

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B12 addition does not alter the dose calculation