rabbit polyclonal anti py397 fak  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: FAK activity exacerbates disturbed flow-mediated atherosclerosis via VEGFR2-Cbl-NF-κB signaling

doi: 10.1101/2024.12.06.627217

Figure Lengend Snippet: HUVECs were starved for 12 h in 0.2% FBS DMEM with or without VEGF (5 ng/ml) prior to initiation of disturbed flow (5 dynes/cm 2 ). (A) Immunoblotting of active FAK (pY397), active NF-κB (pS536), active IKKα/β (pS176/177), IκBα, and β-actin as loading control are shown. (B) HUVECs were treated for 1 h with either DMSO or the FAK inhibitor (FAK-I, 2.5 μM) prior to initiation of disturbed flow (5 dynes/cm 2 ). Immunoblotting shows active FAK (pY397), active NF-κB (pS536), and β-actin as loading control. (C) Immunoblotting of CBL immunoprecipitates (IP) for CBL, FAK, and Tyrosine phosphorylation (4G10). (D) HUVECs were treated with FAK inhibitor (FAK-I, 2.5 μM) prior to initiation of disturbed flow (5 dynes/cm 2 ) for the indicated times. Immunostaining of pY397 FAK (Green) and CBL (Red). Nuclei were stained with DAPI (Blue). (E) HUVECs were treated with FAK inhibitor (FAK-I, 2.5 μM) prior to initiation of disturbed flow (5 dynes/cm 2 ) for the indicated times. Proximity ligation assay (PLA) was performed using antibodies for FAK and CBL. Co- localization is indicated by red dots. Nuclei were stained with DAPI (Blue).

Article Snippet: Anti-pY397 FAK (cat# 44-624G) was purchased from Life Technologies.

Techniques: Western Blot, Control, Immunostaining, Staining, Proximity Ligation Assay

Journal: bioRxiv

Article Title: FAK activity exacerbates disturbed flow-mediated atherosclerosis via VEGFR2-Cbl-NF-κB signaling

doi: 10.1101/2024.12.06.627217

Figure Lengend Snippet: Apoe-/- mice underwent partial carotid ligation (PCL) and were treated with either vehicle or FAK inhibitor (FAK-I, 35 mg/kg) while on a western diet (WD) for 2 weeks. The outer and inner aortic arches were immunostained for (A) active pY397 FAK (Red), (B) Cbl (Red), or (C) Vegfr2 (Red). Endothelial cells were stained with vWF (Green) and nuclei with DAPI (Blue).

Article Snippet: Anti-pY397 FAK (cat# 44-624G) was purchased from Life Technologies.

Techniques: Ligation, Western Blot, Staining

Journal: bioRxiv

Article Title: FAK activity exacerbates disturbed flow-mediated atherosclerosis via VEGFR2-Cbl-NF-κB signaling

doi: 10.1101/2024.12.06.627217

Figure Lengend Snippet: Partial carotid ligation (PCL) was performed on Apoe-/- mice and were treated with vehicle or FAK inhibitor (FAK-I, 30 mg/kg, twice daily) while on a western diet (WD) for 2 weeks. (A) Representative images of Oil Red O staining of sham and ligated carotid arteries are shown. Immunostaining of carotid arteries for (B) pY397 FAK (Red), (C) Cbl (Red), or (D) Vegfr2 (Red). Endothelial cells were stained with Vwf (Green) and nuclei with DAPI (Blue). (E and F) PLA was performed with antibodies targeting (E) FAK and Vegfr2 or (F) FAK and Cbl. Red dots indicate positive co-localization. Nuclei stained with DAPI (blue).

Article Snippet: Anti-pY397 FAK (cat# 44-624G) was purchased from Life Technologies.

Techniques: Ligation, Western Blot, Staining, Immunostaining

Journal: bioRxiv

Article Title: FAK activity exacerbates disturbed flow-mediated atherosclerosis via VEGFR2-Cbl-NF-κB signaling

doi: 10.1101/2024.12.06.627217

Figure Lengend Snippet: Partial carotid ligation (PCL) was performed on Apoe-/-;FAK-WT and Apoe-/-;FAK- KD mice and fed a western diet (WD) for 2 weeks. (A) Representative images of Oil Red O staining of sham and ligated carotid arteries are shown. Immunostaining for (B) pY397 FAK (Red), (C) Cbl (Red), or (D) Vegfr2 (Red). Endothelial cells stained with Vwf (Green) and nuclei were stained with DAPI (blue).

Article Snippet: Anti-pY397 FAK (cat# 44-624G) was purchased from Life Technologies.

Techniques: Ligation, Western Blot, Staining, Immunostaining

Journal: bioRxiv

Article Title: FAK inhibition suppresses breast cancer progression via DNA methylation-mediated DAB2 gene reactivation

doi: 10.1101/2024.11.23.624992

Figure Lengend Snippet: A , MCF7 and MDA231 cells were treated FAK-I (VS-6063, 2.5 μM) for 2 days. Representative immunostaining of Ki-67 (green) and blue (DAPI) were merged. Scale bars, 20 μm. The percentage of Ki-67 positive cells was calculated for each square separately (± SEM, n=3, ** P<0.005 vs. vehicle, unpaired t-test). B , Dot blots for 5-mC using genomic DNA (gDNA) from MCF7 and MDA231 treated with/without FAK-I (2.5 μM) for 2 days. Methylene blue staining of 150 ng total gDNA was used to loading control (n=3). C , Breast cancer cells were treated with FAK-I (VS-6063, 2.5 μM) for 24 h. Shown are immunoblots of lysates for DNMT3A, DNMT3B, DNMT1, pY397 FAK, and GAPDH as loading control (n=3). D , MCF7 and MDA231 cells were treated with FAK-I (2.5 μM) for 24 h. Immunofluorescence staining for DNMT3A (green), DNMT3B (red), and DAPI (blue) are shown (n=3).

Article Snippet: Reagents for immunoblot and immunohistochemistry included FAK (#05-537), 5-mC (SAB2702243), 5-hmC (SAB2702268), Ubiquitin (#04-262), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (#MAB374) antibodies (Millipore, Burlington, MA); DNMT3A (#SC365769), DNMT1 (#SC20701), and DAB2 (#SC136964) antibodies (Santa Cruz Biotech, Dallas, TX); pY397 FAK (#44-624G), and DNMT3B (#PA1-32317) antibodies (Thermo Fisher, Waltham, MA); MG132 and the small molecule FAK inhibitor VS-6063 (MedKoo, Morrisville, NC).

Techniques: Immunostaining, Staining, Control, Western Blot, Immunofluorescence

Journal: bioRxiv

Article Title: FAK inhibition suppresses breast cancer progression via DNA methylation-mediated DAB2 gene reactivation

doi: 10.1101/2024.11.23.624992

Figure Lengend Snippet: A , 4T1 cells treated with FAK-I (2.5 μM) for indicated time. Shown are immunoblots of lysates for pY397 FAK, FAK, Dnmt3a, Dnmt3b, Dnmt1, and Gapdh as loading control (n=3). B , mRNA levels of Dnmnt s were measured after 1 day FAK-I (2.5 μM) treatment (±SEM, n=3). C-E , 4T1 cells were treated with FAK-I (2.5 μM) only or together with MG132 (20 μM) for 12 h. Whole cell lysates were immunoprecipitated with anti-FAK or anti-DNMT3A antibody and subjected to immunoblotting with indicated antibodies (n=3).

Article Snippet: Reagents for immunoblot and immunohistochemistry included FAK (#05-537), 5-mC (SAB2702243), 5-hmC (SAB2702268), Ubiquitin (#04-262), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (#MAB374) antibodies (Millipore, Burlington, MA); DNMT3A (#SC365769), DNMT1 (#SC20701), and DAB2 (#SC136964) antibodies (Santa Cruz Biotech, Dallas, TX); pY397 FAK (#44-624G), and DNMT3B (#PA1-32317) antibodies (Thermo Fisher, Waltham, MA); MG132 and the small molecule FAK inhibitor VS-6063 (MedKoo, Morrisville, NC).

Techniques: Western Blot, Control, Immunoprecipitation

Journal: Cancer Gene Therapy

Article Title: Atypical cellular responses mediated by intracellular constitutive active TrkB (NTRK2) kinase domains and a solely intracellular NTRK2-fusion oncogene

doi: 10.1038/s41417-024-00809-0

Figure Lengend Snippet: Key resources table.

Article Snippet: Rabbit polyclonal anti- pY397-FAK (1:500) , Cell Signaling , Cat# 3283; RRID: AB_2173659.

Techniques: Western Blot, Recombinant, Staining, Membrane, Bicinchoninic Acid Protein Assay, Gel Extraction, Mutagenesis, Software