Blank control: Hela. Primary Antibody (green line): Rabbit Anti-C9orf72 antibody (orb1069), Dilution: 1 µg/10 6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody: Goat anti-rabbit IgG-FITC, Dilution: 1 µg/Test. Protocol, The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 0.1% PBST for 20 min at room temperature. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20000 events was performed.
Sample: 293T (Human) Cell Lysate at 40 ug, Primary: Anti-C9orf72 (orb1069) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 53 kD, Observed band size: 53 kD.
Sample: Kidney (Rat) Lysate at 40 ug, Primary: Anti-C9orf72 (orb1069) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 53 kD, Observed band size: 53 kD.
Sample: Lane 1: Mouse Cerebrum tissue lysates, Lane 2: Mouse Spinal cord tissue lysates, Lane 3: Mouse Cerebellum tissue lysates, Lane 4: Rat Spinal cord tissue lysates, Primary: Anti-C9orf72 (orb1069) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 53 kDa, Observed band size: 50 kDa.
Tissue/Cell: human liver carcinoma, 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-Arginase 1 Polyclonal Antibody, Unconjugated 1:200, overnight at 4C, followed by conjugation to the secondary antibody and DAB staining.
U-937 cells were fixed with 4% PFA for 10 min at room temperature, permeabilized with 90% ice-cold methanol for 20 min at room temperature, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with C9orf72 Antibody (orb1069) at 1:100 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2% BSA in PBS, followed by secondary antibody incubation for 40 min at room temperature. Acquisitions of 20000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).
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