Anti tyrosine hydroxylase

Primary antibodies: α-syn monoclonal mouse-anti-α-syn (SYN-1, BD Biosciences, catalog no. 610787), polyclonal rabbit-anti-α-syn (ASY1^{47 (link)}), rabbit-anti-α-syn (phospho S129) (abcam, catalog no. ab51253), rabbit-anti-α-syn (phospho S129) (Cell Signaling, D1R1R, mAb #23,706), MJFR-14-6-4-2 (MJF14) (abcam, catalog no. 209538, rabbit monoclonal antibody, generated against full length α-syn protein filament), anti-α-tubulin (T9026 Sigma-Aldrich Co.) and anti-Tyrosine Hydroxylase (GeneTex, GTX113016). Secondary antibodies: horseradish peroxidase (HRP)-conjugated polyclonal rabbit-anti-mouse (Dako, P0260), HRP-conjugated polyclonal swine-anti-rabbit (Dako, P0217).

Selected sections were processed for tyrosine hydroxylase immunohistochemistry using the streptavidin–biotin method (EnVision, Dako, Denmark) with diaminobenzidine (DAB) as the electron donor. The antiserum (anti-tyrosine hydroxylase, GeneTex) was diluted in Tris buffer, pH 7.6, containing 0.7% non-gelling seaweed lambda carrageenan (Sigma) and 0.5% Triton X-100 (Sigma). The antiserum was used at a dilution of 1:300. The conditions and duration of incubation with the various reagents, especially with DAB and H₂O₂, was the same in all cases. Use of preimmune serum and omission of incubation in the primary antiserum during the immunostaining procedure were used as test controls and resulted in no immunostaining.