Gibco amniomax c100 basal medium

Explants of ~3-mm dermal biopsies were minced and placed in a 100-mm TC-treated tissue culture dish (Corning Life Science, USA) with a small drop of Fibroblast medium [Gibco AmnioMAX-C100 Basal Medium supplemented with 15% AmnioMAX-C100 Supplement (Invitrogen, Carlsbad, CA, USA). Dishes were incubated at 37°C in a humidified 5% CO₂ atmosphere and media was replaced every 3–4 days. Fibroblast outgrowths were harvested by trypsinization, transferred into a new dish with a modified Fibroblast Medium (1:1 solution of Gibco AmnioMAX-C100 Supplemented with 15% AmnioMAX-C100 Supplement (Invitrogen, Carlsbad, CA, USA) and RPMI-1640 Basal Medium supplemented with 10% fetal bovine serum (Invitrogen), 1X Primocin (Invitrogen), 1% non-essential amino acids (Invitrogen), with media exchange every 3–4 days and allowed to reach 90% confluence prior to splitting. Fibroblast cultures were passaged no more than 3 times prior to collection for DNA, RNA, or protein extracts isolation or cryopreservation.