Anti th polyclonal antibody

Quantification of TH neurons were done based on a previously published protocol^{46 (link)}. Dopaminergic neurons stained with the anti-TH polyclonal antibody (Millipore, 1:1000) were counted manually. Serial sections were cut at 10 μm intervals and slides containing the SN were delineated. Every 10th section was used for manual counting, whereby the number of TH neurons in the sections were averaged as representative of this particular area. The final count is a summation from each mouse and then averaged for each cohort.

TH-positive neurons in the substantia nigra (SN) were counted by staining paraffin-embedded sections with an anti-TH polyclonal antibody (Millipore, Billerica, MA). Stereological counting of DA neurons was performed by counting and tallying TH-immunopositive neurons on every tenth section based on a previously published report [35 (link)].

The animals were anesthetized with ketamine and xylazine (2.6 and 0.16 mg/kg body weight respectively) 8 days after MPTP intoxication. The brain sections were processed as previously explained [10 (link)] and incubated overnight with anti-TH polyclonal antibody (Millipore 1/500 v/v), anti-DAT antibody (Santa Cruz Biotechnology sc14002, 1/300 v/v), anti-COX-2 antibody (Cell signalling, 1/250 v/v), anti-iNOS antibody (BD Transduction, 1/500 v/v), anti-Bax antibody (Santa Cruz Biotechnology, 1/100 in PBS, v/v) and anti-Bcl-2 antibody (Santa Cruz Biotechnology, 1/500 v/v). Immunohistochemical images were collected using a Zeiss microscope and Axio Vision software. For graphic display of densitometric analyses, the intensity of positive staining (brown staining) was measured by computer-assisted color image analysis (Leica QWin V3, UK). The percentage area of immunoreactivity (determined by the number of positive pixels) was expressed as per cent of total tissue area (red staining). Replicates for every experimental condition and histochemical staining were acquired from each mouse in all experimental group.