Top

Vinculin

Cat Number: MAB-80145
Conjugate: Unconjugated
Size: 100 ug
Clone: VIN-54
Concentration: 1mg/ml
Host: Ms
Isotype: IgG1
Immunogen: Human vinculin, purified from uterus
Reactivity: Hu, Ms, Rt, Ch
Applications:

Western Blot: 1:1000 – 1:10.000
with the appropriate system to detect vinculin in cells and tissues.
Immunohistochemistry (Frozen Tissues): 1:50 – 1:200 to detect vinculin in formalin or acetone fixed tissues.
Immunohistochemistry (paraffin-embedded tissues): 1:50-1:200
Immunofluorescence: 1:50-1:200
Flow Cytometry: Optimal dilutions should be determined by end user

Molecular: Calculated Molecular Weight: ~117 kDA Observed Molecular Weight: ~124 kDA
Purification: Purified by the goat anti-mouse IgG affinity chromatography.
Background:

Vinculin is a cytoskeletal protein associated with the cytoplasmic face of both cell-cell and cell-extracellular matrix adherens-type junctions, where it is thought to function as one of several interacting proteins involved in anchoring F-actin to the membrane, both human and chicken embryo sequences of vinculin contain 1,066 amino acids and, furthermore, that the 2 proteins exhibit a high level of sequence identity (greater than 95%). Vinculinis mapped to 10q22.1-q23.

Form: Liquid
Buffer: PBS with 0.02% sodium azide and 50% glycerol pH 7.3
Storage: At 2-8°C for short term storage, and at -20°C for longer term. Avoid freeze and thaw cycles

WB on different cell lysates using Anti-Vinculin MAB-80145 at a dilution of 1:2,000
each lane was loaded with 50 ug of cell lysate
1. human Hela
2. human HepG2
3. monkey COS-7
4. human U-87MG
5. rat PC-12
6. rat RH35
7. mouse HEPA1-6
8. mouse NIH3T3

Anti- Vinculin antibody, IHC(P)Human Placenta Tissue

Anti- Vinculin antibody, IHC(P)Human Placenta Tissue

IHC (Paraffin embedded) PROTOCOL .

Vinculin was detected in paraffin embedded section of human placenta 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-Vinculin Antibody 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) with DAB as the chromogen.

Western Blot protocol with Tanon 5200 system

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 50ug of sample under reducing conditions.

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-Vinculin antigen affinity purified Monoclonal antibody (Cat.# MAB-80145) at 1:2000 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 1:10,000 for 1.5 hour at RT.

The signal is developed using an Enhanced Chemiluminescent detection ECL West Pico kit with Tanon 5200 system. A specific band was detected for Vinculin at approximately 124KD. The expected band size for Vinculin is at 124KD.