For ultrathin cryosectioning, melanocytes were fixed with 2% PFA or with a mixture of 2% PFA and 0.2% glutaraldehyde in 0.1 M phosphate buffer, pH 7.4. Cells were processed for ultracryomicrotomy (70 nm of thickness) and stained using the method described for ultrathin cryosections47 (link). All samples were analysed using a FEI CM120 electron microscope (FEI Company), and digital acquisitions were made with a numeric camera (Keen View; Soft Imaging System, SIS, Germany).
Lo Cicero A., Saidani M., Allouche J., Egesipe A.L., Hoch L., Bruge C., Sigaudy S., De Sandre-Giovannoli A., Levy N., Baldeschi C, & Nissan X. (2018). Pathological modelling of pigmentation disorders associated with Hutchinson-Gilford Progeria Syndrome (HGPS) revealed an impaired melanogenesis pathway in iPS-derived melanocytes. Scientific Reports, 8, 9112.
Type of fixation: 2% PFA or a mixture of 2% PFA and 0.2% glutaraldehyde in 0.1 M phosphate buffer, pH 7.4
dependent variables
Ultrastructural characteristics of melanocytes observed using electron microscopy
control variables
Phosphate buffer, pH 7.4
Ultracryomicrotomy settings (70 nm thickness)
Staining method for ultrathin cryosections as described in reference 47
FEI CM120 electron microscope used for analysis
Keen View numeric camera used for digital acquisitions
controls
No positive or negative controls were explicitly mentioned in the provided information.
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