Cyclophilin E/PPIE Antibody (monoclonal, 4I9)

• Catalog Number: orb763198
• Category: Antibodies
• Description: Cyclophilin E/PPIE Antibody (monoclonal, 4I9)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 4I9
• Tested applications: FC, ICC, IF, IHC, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Mouse IgG2b
• Immunogen: E.coli-derived human Cyclophilin E/PPIE recombinant protein (Position: M1-V301).
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.25-0.5 μg/ml, Human, Mouse, Rat Immunohistochemistry(Paraffin-embedded Section), 2-5 μg/ml, Human, Rat Immunocytochemistry/Immunofluorescence, 5 μg/ml, Human Flow Cytometry, 1-3 μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 35 kDa
• UniProt ID: Q9UNP9
• Storage: At -20°C for one year from date of receipt. After reconstitution, at 4°C for one month. It can also be aliquotted and stored frozen at -20°C for six months. Avoid repeated freezing and thawing.
• Note: For research use only
• Application notes: Tested Species: In-house tested species with positive results. Other applications have not been tested. Optimal dilutions should be determined by end users. Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). 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 HEK293 whole cell lysates, Lane 2: human K562 whole cell lysates, Lane 3: human PC-3 whole cell lysates, Lane 4: human Caco-2 whole cell 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 mouse anti-Cyclophilin E/PPIE antigen affinity purified monoclonal antibody (Catalog # orb763198) 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-mouse IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # orb90502) with Tanon 5200 system. A specific band was detected for Cyclophilin E/PPIE at approximately 35 kDa. The expected band size for Cyclophilin E/PPIE is at 35 kDa.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human breast 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human lung 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of rat heart 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human placenta 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human gall bladder adenosquamous carcinoma 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human liver 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IHC analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in a paraffin-embedded section of human colonic adenocarcinoma 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 2 μg/ml mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. Biotinylated goat anti-mouse 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 # orb90443) with DAB as the chromogen.

IF analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198). Cyclophilin E/PPIE was detected in an immunocytochemical section of HeLa cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (orb90553) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL mouse anti-Cyclophilin E/PPIE Antibody (orb763198) overnight at 4°C. DyLight?594 Conjugated Goat Anti-Mouse IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The tissue section was developed using Phalloidin-iFluor 488 Conjugated. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Flow Cytometry analysis of U937 cells using anti-Cyclophilin E/PPIE antibody (orb763198). Overlay histogram showing U937 cells stained with orb763198 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-Cyclophilin E/PPIE Antibody (orb763198, 1 μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (5-10 μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1 μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

Western blot analysis of Cyclophilin E/PPIE using anti-Cyclophilin E/PPIE antibody (orb763198).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: rat heart tissue lysates, Lane 2: rat brain tissue lysates, Lane 3: rat lung tissue lysates, Lane 4: mouse heart tissue lysates, Lane 5: mouse brain tissue lysates, Lane 6: mouse lung tissue lysates, Lane 7: rat RH35 whole cell lysates, Lane 8: mouse HEPA1-6 whole cell 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 mouse anti-Cyclophilin E/PPIE antigen affinity purified monoclonal antibody (Catalog # orb763198) 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-mouse IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # orb90502) with Tanon 5200 system. A specific band was detected for Cyclophilin E/PPIE at approximately 35 kDa. The expected band size for Cyclophilin E/PPIE is at 35 kDa.