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Protein cytotoxicity assays. To check the putative cytotoxic effects
of transferrins or hOtrf peptides, CEF cells were grown to 70% confluency
in 96-well plates and were exposed to increasing concentrations
of compounds in 2% FCS MEM, from 6 h up to 48 h at 37 C. After
Trypan blue staining, viable cells were counted and cell-associated acid
phosphatase was dosed using p-nitrophenylphosphate as substrate,
according to the Tox-3 kit protocol (Sigma Chemical).
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