Semaphorin 3B/SEMA3B Antibody (monoclonal, 9C4F7)

• Catalog Number: orb865672
• Category: Antibodies
• Description: Semaphorin 3B/SEMA3B Antibody (monoclonal, 9C4F7)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 9C4F7
• Tested applications: FC, ICC, IF, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Mouse IgG2b
• Immunogen: E.coli-derived human Semaphorin 3B/SEMA3B recombinant protein (Position: Y55-R624).
• 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 Immunocytochemistry/Immunofluorescence, 5 μg/ml, Human Flow Cytometry, 1-3 μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 83 kDa
• UniProt ID: Q13214
• 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 Semaphorin 3B/SEMA3B using anti-Semaphorin 3B/SEMA3B antibody (orb865672). 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 CACO-2 whole cell lysates, Lane 3: human U87 whole cell lysates, Lane 4: human K562 whole cell lysates, Lane 5: rat kidney tissue lysates, Lane 6: rat NRK whole cell lysates, Lane 7: mouse kidney tissue lysates, Lane 8: mouse ANA-1 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-Semaphorin 3B/SEMA3B antigen affinity purified monoclonal antibody (Catalog # orb865672) 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 Semaphorin 3B/SEMA3B at approximately 83 kDa. The expected band size for Semaphorin 3B/SEMA3B is at 83 kDa.

IF analysis of Semaphorin 3B/SEMA3B using anti-Semaphorin 3B/SEMA3B antibody (orb865672). Semaphorin 3B/SEMA3B 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-Semaphorin 3B/SEMA3B Antibody (orb865672) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Mouse IgG 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.

Flow Cytometry analysis of HepG2 cells using anti-Semaphorin 3B/SEMA3B antibody (orb865672). Overlay histogram showing HepG2 cells stained with orb865672 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-Semaphorin 3B/SEMA3B Antibody (orb865672, 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.