rabbit polyclonal antibodies against total map1b  (Cell Signaling Technology Inc)

Journal: PLoS ONE

Article Title: Repulsive Axon Guidance by Draxin Is Mediated by Protein Kinase B (Akt), Glycogen Synthase Kinase-3β (GSK-3β) and Microtubule-Associated Protein 1B

doi: 10.1371/journal.pone.0119524

Figure Lengend Snippet: A. Cortical explants from newborn wild-type or MAP1B-/- mice, cultured for 48 h in the presence or absence (control) of 10 nM draxin or 100 ng/ml semaphorin 3A (Sema3A). Scale bar = 100 μm. B. Quantification of the longest neurites of 30 explants in 3 independent experiments. C. Growth cones of cortical neurons from newborn wild-type or MAP1B-/- mice cultured for 60 h, treated for 1 h with 100 nM draxin or 30 min with 100 ng/ml semaphorin 3A (Sema3A), fixed and stained for F-actin. Scale bar = 10 μm. D. Percentage of collapsed growth cones. For each experimental condition the growth cones of 30 neurons in each of 3 independent experiments were evaluated.

Article Snippet: Primary antibodies: mouse monoclonal antibodies against phospho-MAP1B (SMI31; 1:1000; Covance) and neurofilament H (1:1000; Cell Signaling); rabbit polyclonal antibodies against total-MAP1B raised against peptides ATVVVEATEPEPSGC and ETVTEEHLRRAIGN [ ], phospho Ser473 Akt (1:1000; Cell Signaling), total Akt (1:1000; Cell Signaling) and GAPDH (1:3000; Sigma-Aldrich); rabbit monoclonal antibodies against phospho Ser9-GSK-3β (1:1000; Cell Signaling) and total GSK-3β (1:1000; Cell Signaling).

Techniques: Cell Culture, Control, Staining

Journal: PLoS ONE

Article Title: Repulsive Axon Guidance by Draxin Is Mediated by Protein Kinase B (Akt), Glycogen Synthase Kinase-3β (GSK-3β) and Microtubule-Associated Protein 1B

doi: 10.1371/journal.pone.0119524

Figure Lengend Snippet: Immunoblot analyses of cortical neurons from newborn wild-type mice cultured for 60 h, treated with draxin in the absence or presence of the GSK-3β inhibitor SB216763 for the indicated times, lysed and probed using the indicated antibodies. A. Representative immunoblot for determination of MAP1B phosphorylation in response to draxin treatment. B. The relative level of phosphorylated MAP1B (P-MAP1B) was determined by normalizing the P-MAP1B signal to the signal for total MAP1B 30 min after draxin addition in 3 independent experiments. C. Representative immunoblot for determination of MAP1B protein levels in response to draxin treatment. D. The relative level of MAP1B was determined by normalizing the MAP1B signal to the signal for neurofilament H in 3 independent experiments. Draxin treatment did not lead to a significant change in MAP1B levels.

Article Snippet: Primary antibodies: mouse monoclonal antibodies against phospho-MAP1B (SMI31; 1:1000; Covance) and neurofilament H (1:1000; Cell Signaling); rabbit polyclonal antibodies against total-MAP1B raised against peptides ATVVVEATEPEPSGC and ETVTEEHLRRAIGN [ ], phospho Ser473 Akt (1:1000; Cell Signaling), total Akt (1:1000; Cell Signaling) and GAPDH (1:3000; Sigma-Aldrich); rabbit monoclonal antibodies against phospho Ser9-GSK-3β (1:1000; Cell Signaling) and total GSK-3β (1:1000; Cell Signaling).

Techniques: Western Blot, Cell Culture, Phospho-proteomics

Journal: PLoS ONE

Article Title: Repulsive Axon Guidance by Draxin Is Mediated by Protein Kinase B (Akt), Glycogen Synthase Kinase-3β (GSK-3β) and Microtubule-Associated Protein 1B

doi: 10.1371/journal.pone.0119524

Figure Lengend Snippet: Immunoblot analyses of cortical neurons from newborn wild-type ( A ) or MAP1B-/- ( B ) mice cultured for 60 h, treated with draxin for the indicated times, lysed and probed using the indicated antibodies. The GSK-3β doublets represent the GSK-3β1 and GSK-3β2 isoforms .The relative levels of GSK-3β phosphorylated at Ser9 (P-GSK-3β) and Akt phosphorylated on Ser473 (P-Akt) were determined by normalizing the signals for the phosphorylated proteins to the corresponding signals for the total proteins in 3 independent experiments.

Article Snippet: Primary antibodies: mouse monoclonal antibodies against phospho-MAP1B (SMI31; 1:1000; Covance) and neurofilament H (1:1000; Cell Signaling); rabbit polyclonal antibodies against total-MAP1B raised against peptides ATVVVEATEPEPSGC and ETVTEEHLRRAIGN [ ], phospho Ser473 Akt (1:1000; Cell Signaling), total Akt (1:1000; Cell Signaling) and GAPDH (1:3000; Sigma-Aldrich); rabbit monoclonal antibodies against phospho Ser9-GSK-3β (1:1000; Cell Signaling) and total GSK-3β (1:1000; Cell Signaling).

Techniques: Western Blot, Cell Culture

Journal: PLoS ONE

Article Title: Repulsive Axon Guidance by Draxin Is Mediated by Protein Kinase B (Akt), Glycogen Synthase Kinase-3β (GSK-3β) and Microtubule-Associated Protein 1B

doi: 10.1371/journal.pone.0119524

Figure Lengend Snippet: Draxin interaction with DCC triggers inactivation of the Akt pathway. This relieves GSK-3β from Akt-mediated inhibition leading to an increase in phosphorylation of MAP1B and reconfiguration of the cytoskeleton to promote growth cone collapse and inhibition of neurite extension.

Article Snippet: Primary antibodies: mouse monoclonal antibodies against phospho-MAP1B (SMI31; 1:1000; Covance) and neurofilament H (1:1000; Cell Signaling); rabbit polyclonal antibodies against total-MAP1B raised against peptides ATVVVEATEPEPSGC and ETVTEEHLRRAIGN [ ], phospho Ser473 Akt (1:1000; Cell Signaling), total Akt (1:1000; Cell Signaling) and GAPDH (1:3000; Sigma-Aldrich); rabbit monoclonal antibodies against phospho Ser9-GSK-3β (1:1000; Cell Signaling) and total GSK-3β (1:1000; Cell Signaling).

Techniques: Inhibition, Phospho-proteomics