Phosphorylated Akt Detection via Western Blot

Western blots were carried out as previously described [16 (link)]. Immunostaining was carried out using a goat monoclonal antibodies against phospho-Akt (S473) (Cell Signaling #9271) and actin (1/1000, Cell signaling) and a secondary polyclonal mouse anti-goat antibody HRP conjugated (1/2000, cell signaling). Blots were developed using HRP and chemiluminescent peroxidase substrate (#CPS1120, Sigma). Data were collected using Geliance CCD camera (Perkin Elmer), and analyzed using ImageJ software (NIH).

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Hoarau-Véchot J., Touboul C., Halabi N., Blot-Dupin M., Lis R., Abi Khalil C., Rafii S., Rafii A, & Pasquier J. (2019). Akt-activated endothelium promotes ovarian cancer proliferation through notch activation. Journal of Translational Medicine, 17, 194.

Publication 2019

CPS1120 Geliance CCD camera

Actin Anti antibody Goat Monoclonal antibodies Mouse Peroxidase Western blots

Corresponding Organization :

Other organizations : Weill Cornell Medical College in Qatar, Inserm, Hôpital Intercommunal de Créteil, Université Paris-Est Créteil, Cornell University

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Variable analysis

independent variables

Immunostaining using goat monoclonal antibodies against phospho-Akt (S473) and actin

dependent variables

Phospho-Akt (S473) and actin protein levels

control variables

Western blot protocol as previously described [16]
Actin as a loading control
Peroxidase substrate for blot development

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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