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Read our guarantee »Products:Signal Transduction >> Cytoskeleton / ECM >> Cytoskeleton >> Microfilaments >> Actin etc >> Actin Binding Proteins
Anti-Vinculin antibody [SPM227]
See all Vinculin products (14) ...
Mouse monoclonal [SPM227] to Vinculin
ICC/IF, WB, IP, IF, IHC-Pmore details
Reacts with
Mouse, Hamster, Human
Full length native protein (semi-purified) (Human).
HeLa cells, skin.
Liquid
Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
Preservative: 0.09% Sodium Azide
Constituents: BSA, 10mM PBS, pH 7.4
Concentration information loading...
Protein G purified
Purified from ascites fluid by protein G
Monoclonal
SPM227
IgG1
kappa
Cardiovascular >> Heart >> Contractility >> Contractile Proteins >> Actins
Cardiovascular >> Heart >> Cardiac arrhythmias
Signal Transduction >> Cytoskeleton / ECM >> Cytoskeleton >> Microfilaments >> Actin etc >> Actin Binding Proteins
Our Abpromise guarantee covers the use of ab18058 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ICC/IF: 1/200(See Abreview.)
WB: Use a concentration of 1 - 2 µg/ml.Predicted molecular weight: 130 kDa.(130kDa (vinculin) and 150kDa (meta-vinculin))
IP: Use at 2 µg/mg of lysate.
IF: Use at an assay dependent dilution.
IHC-P: 1/25Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Actin filament (F-actin)-binding protein involved in cell-matrix adhesion and cell-cell adhesion. Regulates cell-surface E-cadherin expression and potentiates mechanosensing by the E-cadherin complex. May also play important roles in cell morphology and locomotion.
Metavinculin is muscle-specific.
Defects in VCL are the cause of cardiomyopathy dilated type 1W (CMD1W) [MIM:611407]. Dilated cardiomyopathy is a disorder characterized by ventricular dilation and impaired systolic function, resulting in congestive heart failure and arrhythmia. Patients are at risk of premature death.
Defects in VCL are the cause of cardiomyopathy familial hypertrophic type 15 (CMH15) [MIM:613255]. It is a hereditary heart disorder characterized by ventricular hypertrophy, which is usually asymmetric and often involves the interventricular septum. The symptoms include dyspnea, syncope, collapse, palpitations, and chest pain. They can be readily provoked by exercise. The disorder has inter- and intrafamilial variability ranging from benign to malignant forms with high risk of cardiac failure and sudden cardiac death.
Belongs to the vinculin/alpha-catenin family.
Exists in at least two conformations. When in the closed, 'inactive' conformation, extensive interactions between the head and tail domains prevent detectable binding to most of its ligands. It takes on an 'active' conformation after cooperative and simultaneous binding of two different ligands. This activation involves displacement of the head-tail interactions and leads to a significant accumulation of ternary complexes. The active form then binds a number of proteins that have both signaling and structural roles that are essential for cell adhesion.
The N-terminal globular head (Vh) comprises of subdomains D1-D4. The C-terminal tail (Vt) binds F-actin and cross-links actin filaments into bundles. An intramolecular interaction between Vh and Vt masks the F-actin-binding domain located in Vt. The binding of talin and alpha-actinin to the D1 subdomain of vinculin induces a helical bundle conversion of this subdomain, leading to the disruption of the intramolecular interaction and the exposure of the cryptic F-actin-binding domain of Vt. Vt inhibits actin filament barbed end elongation without affecting the critical concentration of actin assembly.
Phosphorylated; on serines, threonines and tyrosines. Phosphorylation on Tyr-1133 in activated platelets affects head-tail interactions and cell spreading but has no effect on actin binding nor on localization to focal adhesion plaques.
Aceylated; mainly by myristic acid but also small amount of palmitic acid.
Cytoplasm > cytoskeleton. Cell junction > adherens junction. Cell membrane. Cytoplasmic face of adhesion plaques. Recruitment to cell-cell junctions occurs in a myosin II-dependent manner. Interaction with CTNNB1 is necessary for its localization to the cell-cell junctions.
Target information above from: UniProt accessionP18206
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)
![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/Images/18/ab18058/ab18058_1.jpg)
ab18058 at 1/200 staining Chinese Hamster Ovary cells by ICC/IF. The cells were permeabilized with 100% methanol and then blocked with 1% BSA/ 4% goat serum prior to incubation with the antibody for 30 minutes. A rhodamine conjugated goat antibody was used as the secondary.
This image is courtesy of an anonymous Abreview
Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)
![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/Images/18/ab18058/ab18058_2.jpg)
ab18058 staining mouse 3T3 cells by ICC/IF. Cells were PFA fixed and permeabilized in Triton X-100 prior to blocking in 1% BSA for 30 minutes at 25°C. The primary antibody was diluted 1/100 and incubated with the sample for 1 hour at 25°C, ab6785, diluted 1/100, was used as the secondary antibody.
This image is courtesy of an Abreview submitted by Mr John Maloney
Western blot - Vinculin antibody [SPM227] (ab18058)
![Western blot - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-3.jpg)
Anti-Vinculin antibody [SPM227] (ab18058) at 1/1000 dilution + Whole cell lysate prepared from human lung cells at 40 µg
Secondary
HRP conjugated goat polyclonal to mouse IgG at 1/1000 dilution
Predicted band size : 130 kDa
Observed band size : 117 kDa (why is the actual band size different from the predicted?)
The image is a courtsey of an anonymous abreview.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)-Vinculin antibody [SPM227](ab18058)
](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-4.jpg)
Ab18058 staining human normal prostate. Staining is localised to the nucleus and cytoplasm.
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate pH 6.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS), then incubated with primary antibody for 20 minutes, and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
Western blot - Vinculin antibody [SPM227] (ab18058)
![Western blot - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-5.jpg)
Predicted band size : 130 kDa
Lanes: 1, 4, 7 - Hamster Lung; 2, 5, 8 - K562; 3, 6, 9 - CHO
Lanes: 1-3: 4oC (1 freeze/thaw); 4-6: 4oC (2 freeze/thaws); 7-9 : 4oC (4 freeze/thaws)
Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)
![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-8.jpg)
ICC/IF image of ab18058 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab18058, 10µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-mouse IgG - H&L, pre-adsorbed (ab96879) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
This product has been referenced in:
See all 9 publications for this product
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![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/Images/18/ab18058/ab18058_1.jpg)
ab18058 at 1/200 staining Chinese Hamster Ovary cells by ICC/IF. The cells were permeabilized with 100% methanol and then blocked with 1% BSA/ 4% goat serum prior to incubation with the antibody for 30 minutes. A rhodamine conjugated goat antibody was used as the secondary.
This image is courtesy of an anonymous Abreview
![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/Images/18/ab18058/ab18058_2.jpg)
ab18058 staining mouse 3T3 cells by ICC/IF. Cells were PFA fixed and permeabilized in Triton X-100 prior to blocking in 1% BSA for 30 minutes at 25°C. The primary antibody was diluted 1/100 and incubated with the sample for 1 hour at 25°C,
This image is courtesy of an Abreview submitted by Mr John Maloney
![Western blot - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-3.jpg)
Anti-Vinculin antibody [SPM227] (ab18058) at 1/1000 dilution + Whole cell lysate prepared from human lung cells at 40 µg
Secondary
HRP conjugated goat polyclonal to mouse IgG at 1/1000 dilution
Predicted band size : 130 kDa
Observed band size : 117 kDa (why is the actual band size different from the predicted?)
The image is a courtsey of an anonymous abreview.
](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-4.jpg)
Ab18058 staining human normal prostate. Staining is localised to the nucleus and cytoplasm.
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate pH 6.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS), then incubated with primary antibody for 20 minutes, and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
![Western blot - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-5.jpg)
Predicted band size : 130 kDa
Lanes: 1, 4, 7 - Hamster Lung; 2, 5, 8 - K562; 3, 6, 9 - CHO
Lanes: 1-3: 4oC (1 freeze/thaw); 4-6: 4oC (2 freeze/thaws); 7-9 : 4oC (4 freeze/thaws)
![Immunocytochemistry/ Immunofluorescence - Vinculin antibody [SPM227] (ab18058)](/ps/datasheet/images/18/ab18058/Vinculin-Primary-antibodies-ab18058-8.jpg)
ICC/IF image of ab18058 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab18058, 10µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-mouse IgG - H&L, pre-adsorbed (ab96879) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
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