Anti-TCP1 epsilon/CCT5 Antibody Picoband®

Western blot analysis of CCT5 using anti-CCT5 antibody (PB9928).
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 Hela whole cell lysates,
Lane 2: human MCF-7 whole cell lysates,
Lane 3: human SiHa whole cell lysates,
Lane 4: human A549 whole cell lysates,
Lane 5: human 293T whole cell lysates,
Lane 6: human CACO-2 whole cell lysates,
Lane 7: human PC-3 whole cell lysates,
Lane 8: human U20S whole cell lysates,
Lane 9: rat testis tissue lysates,
Lane 10: mouse testis tissue lysates,
Lane 11: mouse liver 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-CCT5 antigen affinity purified polyclonal antibody (Catalog # PB9928) 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-HRP 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 CCT5 at approximately 60 kDa. The expected band size for CCT5 is at 60 kDa.

IHC analysis of CCT5 using anti-CCT5 antibody (PB9928).
CCT5 was detected in paraffin-embedded section of mouse ovary tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-CCT5 Antibody (PB9928) 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.

IHC analysis of CCT5 using anti-CCT5 antibody (PB9928).
CCT5 was detected in paraffin-embedded section of rat ovary tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-CCT5 Antibody (PB9928) 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.

IHC analysis of CCT5 using anti-CCT5 antibody (PB9928).
CCT5 was detected in paraffin-embedded section of human intestinal cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-CCT5 Antibody (PB9928) 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.

IHC analysis of TCP1 epsilon using anti-TCP1 epsilon antibody (PB9928).
TCP1 epsilon was detected in immunocytochemical section of LOVO cell. 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 1μg/ml rabbit anti-TCP1 epsilon Antibody (PB9928) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

IF analysis of CCT5 using anti-CCT5 antibody (PB9928).
CCT5 was detected in immunocytochemical section of U20S cell. 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-CCT5 Antibody (PB9928) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Immunoprecipitating (IP) CCT5 in MCF-7 whole cell lysate.
Western blot analysis of CCT5 using anti-CCT5 antibody (PB9928);
Lane 1: MCF-7 whole cell lysates (30ug);
Lane 2: Rabbit control IgG instead of anti-CCT5 antibody in MCF-7 whole cell lysate;
Lane 3: anti-CCT5 antibody (2μg) + MCF-7 whole cell lysate (500μg).
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-CCT5 antigen affinity purified polyclonal antibody (PB9928) at a dilution of 0.5 μg/mL and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog # BA1054). The signal is developed using ECL Plus Western Blotting Substrate (Catalog # AR1196-200). A specific band was detected for CCT5 at approximately 60 kDa. The expected band size for CCT5 is at 60 kDa.

Flow Cytometry analysis of A431 cells using anti-CCT5 antibody (PB9928).
Overlay histogram showing A431 cells stained with PB9928 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-CCT5 Antibody (PB9928,1μg/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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.