Clathrin heavy chain/CLTC Antibody (monoclonal, 6D3)

• Catalog Number: orb738492
• Category: Antibodies
• Description: Clathrin heavy chain/CLTC Antibody (monoclonal, 6D3)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 6D3
• Tested applications: FC, IHC, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Mouse IgG2b
• Immunogen: E.coli-derived human Clathrin heavy chain/CLTC recombinant protein (Position: R967-Q1668).
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.25-0.5μg/ml, Human, Mouse, Rat Immunohistochemistry (Paraffin-embedded Section), 2-5μg/ml, Human Flow Cytometry, 1-3μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 180 kDa
• UniProt ID: Q00610
• Storage: Store 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 freeze-thaw cycles.
• 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. Add 0.2ml of distilled water will yield a concentration of 500ug/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of Clathrin heavy chain/CLTC using anti-Clathrin heavy chain/CLTC antibody (orb738492). 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 30ug of sample under reducing conditions. Lane 1: human HeLa whole cell lysates, Lane 2: human A431 whole cell lysates, Lane 3: rat brain tissue lysates, Lane 4: mouse brain tissue lysates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA 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-Clathrin heavy chain/CLTC antigen affinity purified monoclonal antibody (Catalog # orb738492) 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 Clathrin heavy chain/CLTC at approximately 180KD. The expected band size for Clathrin heavy chain/CLTC is at 180KD.

IHC analysis of Clathrin heavy chain/CLTC using anti-Clathrin heavy chain/CLTC antibody (orb738492). Clathrin heavy chain/CLTC was detected in paraffin-embedded section of human pancreatic cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2μg/ml mouse anti-Clathrin heavy chain/CLTC Antibody (orb738492) 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 Clathrin heavy chain/CLTC using anti-Clathrin heavy chain/CLTC antibody (orb738492). Clathrin heavy chain/CLTC was detected in paraffin-embedded section of human renal carcinoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2μg/ml mouse anti-Clathrin heavy chain/CLTC Antibody (orb738492) 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 Clathrin heavy chain/CLTC using anti-Clathrin heavy chain/CLTC antibody (orb738492). Clathrin heavy chain/CLTC was detected in paraffin-embedded section of human tonsil tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2μg/ml mouse anti-Clathrin heavy chain/CLTC Antibody (orb738492) 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.

Flow Cytometry analysis of HepG2 cells using anti-Clathrin heavy chain/CLTC antibody (orb738492). Overlay histogram showing HepG2 cells stained with orb738492 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-Clathrin heavy chain/CLTC Antibody (orb738492, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.