Western Blot Analysis of Protein Expression

Total protein extracts, SDS-PAGE separation and Western Blot were performed with standard methods as described elsewhere^{57 (link),58 (link)}. Antibodies were as follows: goat anti-β-actin #SC-1616, mouse anti-α Tubulin TU-02 #SC-8035, rabbit anti-CCND1 #SC-717 and mouse anti-MYCN #SC-53993, mouse anti-Vinculin #SC-73614, mouse anti-Math1 (DSHB), goat anti-DCX # SC-8066 (Santa Cruz Biotechnology); rabbit anti-PARP1 #9542, mouse anti-Gli1 #L42B10, rabbit anti-cleaved caspase-3 #9661 (Cell Signaling Technology Inc); rabbit anti-Myc #C3956 (Sigma Aldrich); rabbit anti-ZIC1 ab72694 (ABCAM), rabbit anti-Pax6 #PRB-278P (BioLegend), mouse anti-Nestin ab11306 (ABCAM). All antibodies were previously used in^{56 (link),59 (link)–63 (link)}. Immunoreactive bands were visualized by enhanced chemoluminscence using WesternBright ECL HRP substrate (Advansta).

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Petroni M., Sahùn Roncero M., Ramponi V., Fabretti F., Nicolis Di Robilant V., Moretti M., Alfano V., Corsi A., De Panfilis S., Giubettini M., Di Giulio S., Capalbo C., Belardinilli F., Coppa A., Sardina F., Colicchia V., Pedretti F., Infante P., Cardinali B., Tessitore A., Canettieri G., De Smaele E, & Giannini G. (2019). SMO-M2 mutation does not support cell-autonomous Hedgehog activity in cerebellar granule cell precursors. Scientific Reports, 9, 19623.

Publication 2019

mouse anti-Gli1 #L42B10 rabbit anti-cleaved caspase-3 #9661 rabbit anti-Pax6 #PRB-278P WesternBright ECL HRP substrate

Actin Antibodies Caspase 3 Ccnd1 Goat Mouse Mycn Nestin Parp1 Protein Rabbit Sds page Vinculin Western blot α tubulin

Corresponding Organization :

Other organizations : Italian Institute of Technology, Sapienza University of Rome, Institute of Cell Biology and Neurobiology, National Research Council, University of L'Aquila, Istituto Pasteur

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Protein expression levels of β-actin, α Tubulin, CCND1, MYCN, Vinculin, Math1, DCX, PARP1, Gli1, cleaved caspase-3, Myc, ZIC1, Pax6, Nestin

control variables

None explicitly mentioned

controls

Positive controls: The antibodies used were previously used in the studies cited (references 56, 59-63).
Negative controls: No negative controls were explicitly mentioned.

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