Ta150041 100

Anti α-L-fucosidase (FUCA1) is a polyclonal antibody from Proteintech Group (Rosemont, IL, USA) (16420-1-AP) which recognizes the α-L-fucosidase-1 protein. Anti-turbo-GFP-tag is a monoclonal antibody from Origene Technologies (TA150041-100) which recognizes over expressed recombinant proteins containing the turbo-GFP tag fused to either the amino- or carboxy-termini of targeted proteins. Monoclonal anti-α-tubulin (#T9026) was from Sigma Aldrich (St Louis, MO, USA). Secondary antibodies coupled to horseradish peroxidase were from Amersham Pharmacia Biotech (Piscataway, NJ, USA). Monoclonal anti-α-tubulin (#T9026) was from Sigma Aldrich (St Louis, MO, USA). Secondary antibodies coupled to horseradish peroxidase were from Amersham Pharmacia Biotech (Piscataway, NJ, USA).

All cell lysates were heated at 70 °C to denature proteins, sonicated to shear genomic DNA and then electrophoresed through Tris-Glycine gels or NuPAGE Bis-Tris gels (Thermo Fisher Scientific) for SDS-PAGE separation by mass. Proteins were transferred to nitrocellulose membranes using the Trans-Blot Turbo Transfer System (Bio-Rad, Hercules, CA, USA). The blocking of membranes and subsequent antibody incubations were performed using Odyssey Blocking Buffer according to the manufacturer’s instructions (LI-COR Biosciences, Lincolin, NE, USA). Primary antibodies against mouse NeuN (12943, Cell Signaling Technologies, Danvers, MA, USA; ab177487, Abcam), β-Tubulin (926–42211, LI-COR Biosciences), β-actin (926–42210, LI-COR Biosciences), FLAG (F1804, Millipore Sigma, Burlington, MA), mCherry (ab167453, Abcam, Cambridge, United Kingdom), GAPDH (ab8245, Abcam) and turboGFP (TA150041-100, OriGene) were obtained from commercial sources. LI-COR’s secondary antibodies, IRDye 800CW-conjugated and IRDye 680-conjugated antibodies, were used based on the manufacture’s protocol. Immunoblot signals were visualized by LI-COR’s Odyssey CLx infrared imaging system and quantified by Image Studio Lite (LI-COR Biosciences).