Journal: Molecular Therapy
Article Title: Tight Long-term Dynamic Doxycycline Responsive Nigrostriatal GDNF Using a Single rAAV Vector
Figure Lengend Snippet: GDNF and VP-16 expression patterns . The GDNF expression pattern of the final ON group ( a , b ) was similar to that previously observed where the GDNF transgene (red) is observed distally from that of the transduced cells and VP-16 (green). ( b ) Increased magnification of area outlined in a . VP-16 expression in the OFF group ( c , d ) was similar to that seen in the ON group; however, no GDNF protein expression was observed. GDNF protein expression in the CBA-GDNF group ( e , f ) extended in a diffuse pattern into the VTA and the DMN indicating that GDNF protein was distributed away from the transduced cells in contrast to the pattern of expression observed from an intracellular transgene such as the SN-GFP group ( g , h ). As expected, no immunoreactivity was observed for VP-16 in either the CBA-GDNF group ( e , f , green) or the SN-GFP group ( g , h , red). ( f and h ) Higher magnifications of areas outlined in e and g , respectively. Bar: a , c , e , g = 1.0 mm, bar: b , d , f , h = 0.1 mm. CBA, chicken β-actin; DMN, deep mesencephalic nucleus; GDNF, glial cell line-derived neurotrophic factor; SNc, substantia nigra pars compacta; SNr, substantia nigra pars reticulate; VTA, ventral tegmental area.
Article Snippet: Primary antibodies used were mouse anti-TH (1:2,000), rabbit anti-GFP (1:2,000), mouse anti-calbindin (1:200), rabbit anti-VP-16 (1:50) (Chemicon, Temecula, CA), goat anti-GDNF (1:1,000) (R & D Systems, Minneapolis, MN).
Techniques: Expressing, Crocin Bleaching Assay, Derivative Assay