Human Phospho-p70 S6 Kinase (T389) Antibody

Detection of Human Phospho-p70 S6 Kinase (T389)/p85 S6 Kinase (T412) by Western Blot. Western blot shows lysates of MCF-7 human breast cancer cell line untreated (-) or treated (+) with 100 ng/mL Recombinant Human IGF-I (Catalog # 291-G1) for 60 minutes. PVDF membrane was probed with 0.1 µg/mL of Rabbit Anti-Human Phospho-p70 S6 Kinase (T389) Monoclonal Antibody (Catalog # MAB8963) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for Phospho-p70 S6 Kinase (T389) and Phospho-p85 S6 Kinase (T412) at approximately 70 and 85 kDa, respectively (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Phospho-p70 S6 Kinase (T389) in MCF‑7 Human Cell Line. p70 S6 Kinase phosphorylated at T389 was detected in immersion fixed serum starved MCF-7 human breast cancer cell line untreated (lower panel) or treated with Recombinant Human IGF-I (Catalog # 291-G1) using Rabbit Anti-Human Phospho-p70 S6 Kinase (T389) Monoclonal Antibody (Catalog # MAB8963) at 2 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Rabbit IgG Secondary Antibody (red; Catalog # NL004) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm and nuclei. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Detection of Mouse p70 S6 Kinase/S6K by Western Blot E2 regulates rapamycin effects on mTORC2 activity.A-H, Rapamycin lowers mTORC1 activity independent of presence of E2, ER alpha - or ER beta -agonist, however lowers mTORC2 activity dependent on presence of E2. A-D, HL-1 cells were grown to near confluence in medium containing 10 nM E2, and E-H, 10 nM ER alpha -agonist PPT or 1 nM ER beta -agonist DPN and serum starved for 24 hours prior to incubation with 20 nM rapamycin and IGF-1 for 24 h. A and E show representative westernblots. Equal loading was verified by blotting with antibodies against beta -actin or alpha -tubulin. For B,C,D and F,G,H, labeled bands were quantified with ImageJ software, normalized to loading and IGF-1 induced phosphorylation of indicated proteins was determined by ratio to the value of non-IGF-1 stimulated control cells. Mean ± SEM of fold stimulation by IGF-1 is shown of at least 3 independently performed experiments. * p < 0.05,** p < 0.009, *** p < 0.0001. If not indicated differently, significances are related to respective non-IGF-1 stimulated cells. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0123385), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse p70 S6 Kinase/S6K by Western Blot E2 regulates rapamycin effects on mTORC2 activity.A-H, Rapamycin lowers mTORC1 activity independent of presence of E2, ER alpha - or ER beta -agonist, however lowers mTORC2 activity dependent on presence of E2. A-D, HL-1 cells were grown to near confluence in medium containing 10 nM E2, and E-H, 10 nM ER alpha -agonist PPT or 1 nM ER beta -agonist DPN and serum starved for 24 hours prior to incubation with 20 nM rapamycin and IGF-1 for 24 h. A and E show representative westernblots. Equal loading was verified by blotting with antibodies against beta -actin or alpha -tubulin. For B,C,D and F,G,H, labeled bands were quantified with ImageJ software, normalized to loading and IGF-1 induced phosphorylation of indicated proteins was determined by ratio to the value of non-IGF-1 stimulated control cells. Mean ± SEM of fold stimulation by IGF-1 is shown of at least 3 independently performed experiments. * p < 0.05,** p < 0.009, *** p < 0.0001. If not indicated differently, significances are related to respective non-IGF-1 stimulated cells. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0123385), licensed under a CC-BY license. Not internally tested by R&D Systems.