Tsika E, Moysidou M, Guo J, Cushman M, Gannon P, Sandaltzopoulos R, Giasson BI, Krainc D, Ischiropoulos H, Mazzulli JR. Distinct region-specific alpha-synuclein oligomers in A53T transgenic mice: implications for neurodegeneration. J Neurosci. 2010 Mar 3;30(9):3409-18. PubMed Abstract

REAGENTS/MATERIAL:
Immunohistochemistry: Brain sections from symptomatic A53T+/+ α-syn transgenic mice and age-matched nTg controls were stained with mouse monoclonal antibody syn505 specific for human α-syn. Adjacent sections were stained with rabbit anti-GFAP (Abcam).
Western blotting:. For Western blot analysis of samples sequentially extracted, the following primary antibodies were used: mouse monoclonal anti-α-syn (LB509); mouse monoclonal anti-tyrosine hydroxylase (6D7) (EMD Biosciences); rabbit anti-neuronal specific enolase (Polysciences) and mouse monoclonal anti-vimentin (RV202) (BD Pharmingen).
For Western blot analysis of fractions from gel filtration profiles, the following antibodies were used: mouse monoclonal anti-α-syn (syn211) and rabbit anti-NSE (Polysciences). Fractionated lysates were also blotted with mouse monoclonal anti-α-syn (syn208) and anti-α-syn (syn303). Dopaminergic markers were detected with mouse monoclonal anti-L-3,4-dihydroxyphenylalanine decarboxylase (Alpha Diagnostic) and anti-dopamine transporter (Millipore Bioscience Research Reagents).
Immunodepletion: To immunodeplete α-syn oligomers for control samples, mouse monoclonal anti-α-syn (syn211) (Sigma) was incubated with 100 μg of fractionated lysate.
Assessment of neurodegeneration: Primary cortical neurons were incubated with anti-neurofilament (2H3) (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA) and scanned.