Poly d lysine

Fertilized chicken eggs were incubated at 37 °C and 80% humidity for 8 days. DRG were isolated from the trunk region of an 8-day old chick embryo based on protocols described elsewhere [35 (link),36 ]. In the case of DRG single-cell culture, DRG were dissociated enzymatically by incubation in 0.25% trypsin/0.05% EDTA solution for 20 min and trituration through a small-bore pipette (20–200 µL) [36 ]. DRG explants or cells were cultured in N2 medium [98% DMEM/F-12 medium, 0.5 mg/mL bovine serum albumin (BSA), 2 mM l-glutamine, 1 × N-2 supplement, 100 ng/mL nerve growth factor (NGF)] [36 ]. DRG explants or cells were seeded on coated glass coverslips. The coverslips were coated first with poly-D-lysine (0.1 mg/mL) (Santa Cruz Biotechnology Inc., Heidelberg, Germany) for 1 h at room temperature, and after washing with sterile water, they were coated with laminin from Engelbreth-Holm-Swarm murine sarcoma basement membrane (EHS laminin, laminin-1) (Merck, Darmstadt, Germany) for 4 h in 37 °C. Laminin was used in the concentration of 1 µg/cm² in Hanks’ Balanced Salt solution (HBSS). DRG were cultured at 37 °C in 5% CO₂ for 48 h. Next, they were subjected to lysates collection or fixation for immunostaining.