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21李東木蟲 (職業(yè)作家)
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Preparations used for ophthalmic purpose need to show minimal cytotoxicity. In order to meet this requirement, preparations were tested using keratinocyte epithelial cells (HaCaT). HaCaT cells are a valuable alternative to primary epithelial cell lines in order to assess whether the nanoparticles would affect viability of cells in general . The evaluation of the in vitro cytotoxicity of the NPs was based on cell viability, determined by the MTT assay. In brief, cells were cultured in RPMI 1640 GlutaMax (Invitrogen) supplemented with 10% foetal bovines serum (Invitrogen) and 100 lg/ml Penicillin/100 U Streptomycin (Invitrogen) . Nanoparticle preparations (1 ll) were diluted in growth medium,and cells were further cultured for 24-h incubation at 37 C/5% CO2. A 0.2% (w/V) benzalkonium chloride solution (BKC) was used as positive control for cytotoxicity. The amount of surviving cells was quantified by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis as described by Dillen et al. Additional experiments with undiluted formulations were performed in order to mimic the in vivo situation on the corneal surface. Cells were incubated for 10 min with a dilution series of formulation 6 and 8, and cytotoxicity was evaluated as discussed above. |
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