insr β primary antibody  (Cell Signaling Technology Inc)

Journal: Endocrinology

Article Title: IGF1 Signaling Regulates Neuropeptide Expression in Hypothalamic Neurons Under Physiological and Pathological Conditions

doi: 10.1210/endocr/bqaf051

Figure Lengend Snippet: List of antibodies for Western blots

Article Snippet: INSR-β Primary Antibody , Cell Signaling Technology , 3025 RRID:AB_2280448.

Techniques: Western Blot

Journal: Oncotarget

Article Title: Compounds from the marine sponge Cribrochalina vasculum offer a way to target IGF-1R mediated signaling in tumor cells

doi: 10.18632/oncotarget.10361

Figure Lengend Snippet: A. Western blot analyses of InsR α and InsR β in untreated diploid fibroblasts WI-38, NSCLC U-1810, breast cancer MDA-MB-231 and ovarian carcinoma A2780 or SKOV-3. β-Tubulin was used to visualize equal loading. Resulting bands were scanned by densitometry and fold to DMSO-treated cells is given after correction for loading differences. B. NSCLC U-1810 were treated with IC 50 of compound 1 (1.5 μmol/L) or 2 (15.1 μmol/L) for 4, 8, 16 and 24 hours or with equal volume of DMSO for 24 hours and InsR α and InsR β expression were examined. β-Tubulin was used to visualize equal loading. Resulting bands were scanned by densitometry and fold to DMSO-treated cells is given after correction for loading differences. C. NSCLC U-1810 cells were exposed to IC 50 of compound 1 (1.5 μmol/L). Slides were stained with two different InsR β antibodies in PLA (red) and DAPI (blue) was applied to visualize cell nuclei. Scale bars, 50 μm.

Article Snippet: Two primary antibodies against InsR β from different species (Rabbit: InsR-β, Cell signaling; 3025 and Mouse: InsR β, Abcam; AB69508) were applied.

Techniques: Western Blot, Expressing, Staining

Journal: Molecular Metabolism

Article Title: Inter-domain tagging implicates caveolin-1 in insulin receptor trafficking and Erk signaling bias in pancreatic beta-cells

doi: 10.1016/j.molmet.2016.01.009

Figure Lengend Snippet: InsR-TagRFP is transported to lysosomes in Cav-1 and Flot-1 positive vesicles . ( A ) Confocal imaging of Cav1-mRFP and InsRA-TagBFP in MIN6 cells (n = 10). Scale bars = 10 μm. ( B ) G-STED microscopy in MIN6 cells illustrates the presence of Flot1 on vesicles containing endogenous insulin receptors. Scale bars = 5 μm. ( B′ ) Insets show flotillin-1 positive vesicles harboring insulin receptors. ( C–G ) Colocalization between InsRA-TagBFP and Flot1-mRFP, Flot1-mRFP and CAV1-eGFP, Flot1-mRFP and LAMP2-eGFP, InsRA-TagRFP and LAMP2-eGFP, CAV1-mRFP and LAMP2-eGFP in MIN6 cells demonstrates a Cav1-Flot1-LAMP2 endocytic trafficking route to lysosomes (n = 10). Scale bars = 10 μm; F: Scale bar = 5 μm. ( H, I ) Immunolabeling of endogenous insulin receptors, endogenous Flot1, and endogenous LAMP2 (lysosomes) in primary 12 week-old mouse islet cells (n = 10). Scale bars = 10 μm. ( J ) Summary of object-based colocalization of tagged proteins compared with InsRA-TagRFP in MIN6 cells. Note that InsRA controls represent the maximum colocalization in this system (n = 10 cells per condition). ( K ) Summary of colocalization within vesicular pools in the Cav1/Flot1/Lamp2 pathway of MIN6 cells (n = 10 cells per condition). ( A–I ) MIN6 cells were cultured in 25 mM glucose and primary cells were cultured in 11 mM glucose prior fixation.

Article Snippet: For immunofluorescence analysis, cells were fixed in 4% (w/v) paraformaldehyde (Sigma–Aldrich) for 20 min and permeabilized subsequently in 0.1% (v/v) Triton X-100 (Sigma–Aldrich) for 15 min. Primary antibodies targeting InsR (Cat. #3020S), Clathrin (Cat. # 4796), Cav1 (Cat. #3267), Rab7 (Cat. #9367), and Igf1r (Cat. #3027) from Cell Signalling (Danvers, MA), Flotillin-1 (Cat. #3868-1) from Epitomics (Burlingame, CA), Rab4a (Cat. #sc-26562), Rab5a (Cat. # sc-166600) from Santa Cruz Biotechnology (Dallas, TX), LAMP2 (Cat. #PRS3627) from Sigma–Aldrich (St. Louis, MO), and Na/K pump (Cat. # ab76020) from Abcam (Eugene, OR) were applied for 12 h at 4 °C.

Techniques: Imaging, Microscopy, Immunolabeling, Cell Culture