Anti tyrosinated α tubulin

Western blotting was performed according to standard techniques. Briefly, we subjected 2–10 μg of S2 cell lysates to SDS-PAGE analysis (12.5% for Histone H3; for others, 7.5% or 10%) and immunoblotting. The following antibodies were used for immunoblotting: the anti-Strip antibody14 (link) (rabbit, 1:200), the anti-Strip antibody14 (link) (rat, 1:50), the anti-α-tubulin (mouse, 1:2000, Sigma T6199), the anti-acetylated-α-tubulin (mouse, 1:1000, Sigma T7451), the anti-tyrosinated-α-tubulin (mouse, 1:1000, Sigma T9028), the anti-Histone H3 antibody (rabbit, 1:2000, Active motif 39163) and the anti-TBCD antibody16 (link) (guinea pig, 1:1000). All data are representative of more than three biological replicates.

Immunohistochemistry was performed as described previously [5 (link)]. For this study, we used combined stainings of anti-α-tubulin or anti-serotonin with phalloidin-rhodamine. For anti-α-tubulin staining, a mixture of anti-acetylated α-tubulin (monoclonal anti-tubulin, acetylated antibody, produced in mouse, ascites fluid, Sigma-Aldrich, St. Louis, MO, USA; dilution 1:500 in PBST-NGS) and anti-tyrosinated α-tubulin (monoclonal anti-tubulin, tyrosine antibody, produced in mouse, ascites fluid, Sigma-Aldrich, St. Louis, MO, USA; dilution 1:250 in PBST-NGS) was used. The labeled α-tubulin is a structural component of microtubules, which are amongst others present in axons, while serotonin (=5-HT) is a neurotransmitter. Phalloidin-rhodamine labels filamentous muscular actin (f-actin) [5 (link)].