Anti-Flotillin 2/FLOT2 Antibody Picoband™

Figure 1. Western blot analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human 293T whole cell lysates,
Lane 2: human Hela whole cell lysates,
Lane 3: human A549 whole cell lysates,
Lane 4: rat lung tissue lysates,
Lane 5: mouse lung tissue lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-Flotillin 2 antigen affinity purified polyclonal antibody (Catalog # PB9623) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-DyLight 647 Conjugated secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for Flotillin 2 at approximately 47 kDa. The expected band size for Flotillin 2 is at 47 kDa.

Figure 2. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in a paraffin-embedded section of mouse intestine tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

Figure 3. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in a paraffin-embedded section of rat kidney tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

Figure 4. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in a paraffin-embedded section of human mammary cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

Figure 5. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in frozen section of mouse kidney tissue. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

Figure 6. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in frozen section of rat kidney tissue. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

Figure 7. IHC analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in frozen section of rat small intestine tissue. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

Figure 8. IF analysis of Flotillin 2 using anti-Flotillin 2 antibody (PB9623).
Flotillin 2 was detected in immunocytochemical section of A431 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 2μg/mL rabbit anti-Flotillin 2 Antibody (PB9623) overnight at 4°C. DyLight®550 Conjugated Goat Anti-Rabbit IgG (BA1135) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Figure 9. Flow Cytometry analysis of RH35 cells using anti-Flotillin 2 antibody (PB9623).
Overlay histogram showing RH35 cells stained with PB9623 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-Flotillin 2 Antibody (PB9623,1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

Figure 10. Flow Cytometry analysis of U20S cells using anti-Flotillin 2 antibody (PB9623).
Overlay histogram showing U20S cells stained with PB9623 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-Flotillin 2 Antibody (PB9623,1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample (Red line) was also used as a control.