Figure 8. Nigral TH phosphorylation stoichiometry in aging.

a. Representative blot showing the 30% decrease in TH phosphorylation at ser31 in the 30-month old BNF rat. All TH phosphorylation assays are conducted against a standard curve with quantified site-specific phosphorylation levels so actual quantities of phosphorylated TH at each phosphorylation site are loaded in the standard curve range and the quantity of phosphorylated TH from the sample is interpolated from the standard curve. Because actual quantities of total TH (tTH) vary from sample-to-sample, it is not always possible to load equal amounts of tTH. Total protein load among samples is kept within 25 µg if at all possible to keep any carrier protein effects upon the signal to an absolute minimum. In this case, total protein load differed by 10 µg (80 µg for the 12 mo sample and 90 µg for the 30 mo sample). The standard curve source is from a calibrated PC12 cell extract with a phosphorylation stoichiometry at ser31 at 0.09 ng p31 per ng tTH. b. Site-specific phosphorylation of TH in the SN in test subjects. The 30-month group had significantly reduced phosphorylation at ser 31 (p = 0.029, unpaired Student's t-test). No significant change was observed at ser40 or ser19.