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Blank control (blue line): K562 (blue). Primary Antibody (green line): Rabbit Anti-GLUT4 antibody (orb10728), Dilution: 1 µg/10 6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody (white blue line): Goat anti-rabbit IgG-PE, Dilution: 1 µg/Test. Protocol, The cells were fixed with 70% ethanol (overnight at 4C) and then permeabilized with 0.1% PBS-Tween for 20 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The cells were then incubated in 1 X PBS/2% BSA/10% goat serum to block non-specific protein-protein interactions followed by the antibody for 15 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20000 events was performed. |
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Sample: Heart (Mouse) Lysate at 40 ug, Primary: Anti-GLUT4 (orb10728) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/10000 dilution, Predicted band size: 54 kD, Observed band size: 54 kD. |
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Sample: Heart (Rat) Cell Lysate at 40 ug, Primary: Anti-GLUT4 (orb10728) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 54 kD, Observed band size: 54 kD. |
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Sample: Lane 1: NIH/3T3 (Mouse) Cell Lysate at 30 ug, Lane 2: Heart (Mouse) Lysate at 40 ug, Lane 3: Heart (Rat) Lysate at 40 ug, Lane 4: Muscle (Mouse) Lysate at 40 ug, Lane 5: Muscle (Rat) Lysate at 40 ug, Primary: Anti-GLUT4 (orb10728) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 51 kD, Observed band size: 51 kD. |
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Tissue/Cell: NIH/3T3 cell, 4% Paraformaldehyde-fixed, Triton X-100 at room temperature for 20 min, Blocking buffer (normal goat serum) at 37C for 20 min, Antibody incubation with (GLUT4) polyclonal Antibody, Unconjugated (orb10728) 1:100, 90 minutes at 37C, followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37C for 90 minutes, DAPI (blue) was used to stain the cell nuclei. |
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Tissue/Cell: rat kidney tissue, 4% Paraformaldehyde-fixed and paraffin-embedded, Antigen retrieval: citrate buffer (0.01M, pH6.0), Boiling bathing for 15 min, Block endogenous peroxidase by 3% Hydrogen peroxide for 30 min, Blocking buffer (normal goat serum) at 37C for 20 min, Incubation: Anti-GLUT4 Polyclonal Antibody, Unconjugated (orb10728) 1:200, overnight at 4C, followed by conjugation to the secondary antibody and DAB staining. |