Anti-Vinculin antibody [EPR20407] KO tested (ab219649)

Anti-Vinculin antibody [EPR20407] ab219649 is a rabbit monoclonal antibody that is used in Vinculin western blotting and IHC. Suitable for human, mouse and rat samples.

- Recombinant format for unrivaled batch-batch consistency: no need for same-lot requests
- Specificity confirmed with VCL knockout cell line validation
- Specificity and sensitivity confirmed in IHC with multi-tissue microarray (TMA) validation

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Images

Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (AB219649), expandable thumbnail

Publications

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	WB	IHC-P
Human	Tested	Tested
Mouse	Tested	Tested
Rat	Tested	Tested

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/1000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Rat	Dilution info 1/1000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Human	Dilution info 1/1000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/2000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Rat	Dilution info 1/2000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Human	Dilution info 1/2000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Associated Products

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Target data

See full target information VCL

Function

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.

Alternative names

Vinculin, Metavinculin, MV, VCL

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Clone number: EPR20407
Purification technique: Affinity purification Protein A
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage duration: 1-2 weeks
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: -20°C
Aliquoting information: Upon delivery aliquot
Storage information: Avoid freeze / thaw cycle

Notes

Patented technology
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

What are the advantages of a recombinant monoclonal antibody?
This product is a recombinant monoclonal antibody, which offers several advantages including:

- High batch-to-batch consistency and reproducibility
- Improved sensitivity and specificity
- Long-term security of supply
- Animal-free batch production

For more information, read more on recombinant antibodies.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

11 product images

Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649)
Vinculin Western blot staining using rabbit Anti-Vinculin antibody
Lanes 1 - 4: Merged signal (red and green). Green - ab219649 observed at 124 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
ab219649 was shown to react with VCL in A431 wild-type cells in Western blot. Loss of signal was observed when VCL knockout sample was used. A431 wild-type and VCL knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% Milk in TBS-T (0.1% Tween®) before incubation with ab219649 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649) at 1/1000 dilution
Lane 1: Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg
Lane 2: VCL knockout A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg
Lane 2: Western blot - Human VCL (Vinculin) knockout A-431 cell line (Human VCL (Vinculin) knockout A-431 cell line ab261898)
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4: Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 124 kDa
Observed band size: 124 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded human breast tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous staining on myoepithelium of human breast is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded human colon tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous and cytoplasmic staining on smooth muscle cells of human colon is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649)
Vinculin Western blot staining using rabbit Anti-Vinculin antibody
Blocking/Dilution buffer: 5% NFDM/TBST.
All lanes: Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649) at 1/1000 dilution
Lane 1: PC-3 (Human prostate adenocarcinoma cell line) whole cell lysate at 20 µg
Lane 2: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 3: K562 (Human chronic myelogenous leukemia cell line from bone marrow) whole cell lysate at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 124 kDa
Observed band size: 123 kDa
Exposure time: 15s
Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649)
Vinculin Western blot staining using rabbit Anti-Vinculin antibody
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure time: Lane 1-3: 15 seconds; Lane 4: 30 seconds.
All lanes: Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649) at 1/1000 dilution
Lane 1: Human fetal kidney lysate at 20 µg
Lane 2: Human fetal heart lysate at 20 µg
Lane 3: Human testis lysate at 20 µg
Lane 4: Human fetal liver lysate at 20 µg
Secondary
All lanes: Western blot - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/4000 dilution
Predicted band size: 124 kDa
Observed band size: 123 kDa
Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649)
Vinculin Western blot staining using rabbit Anti-Vinculin antibody
Blocking/Dilution buffer: 5% NFDM/TBST.
All lanes: Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649) at 1/1000 dilution
Lane 1: Mouse heart lysate at 10 µg
Lane 2: Mouse spleen lysate at 10 µg
Lane 3: Rat heart lysate at 10 µg
Lane 4: Rat kidney lysate at 10 µg
Lane 5: Rat spleen lysate at 10 µg
Lane 6: C6 (Rat glial tumor cell line) whole cell lysate at 10 µg
Lane 7: PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate at 10 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 124 kDa
Observed band size: 123 kDa
Exposure time: 4s
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded human gastric cancer tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous and cytoplasmic staining on smooth muscle cells of human gastric cancer is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded human prostate cancer tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous and cytoplasmic staining on smooth muscle cells of human prostate cancer is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous and cytoplasmic staining on smooth muscle cells of mouse colon is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vinculin antibody [EPR20407] (ab219649)
Immunohistochemical analysis of paraffin-embedded rat stomach tissue labeling Vinculin with ab219649 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Membranous and cytoplasmic staining on smooth muscle cells of rat stomach is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Vinculin antibody [EPR20407] - Loading Control (ab219649)
Vinculin Western blot staining using rabbit Anti-Vinculin antibody
False colour image of Western blot: Anti-PD-L1 antibody [CAL10] – Mouse IgG1 (Chimeric); Rabbit anti-Vinculin antibody (ab219649) loading control staining at 1/1000 dilution, shown in red. In Western blot, Anti-PD-L1 antibody [CAL10] - Mouse IgG1 (Chimeric) ab279292 was shown to bind specifically to PD-L1. A band was observed at 45 kDa in wild-type A549 cell lysates with no signal observed at this size in Cd274 knockout cell line. To generate this image, wild-type and Cd274 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.
All lanes: Western blot - Anti-PD-L1 antibody [CAL10] - Mouse IgG1 (Chimeric) (Anti-PD-L1 antibody [CAL10] - Mouse IgG1 (Chimeric) ab279292) at 1/1000 dilution
Lanes 1 - 2: Western blot - Human wild-type A549 cell line (ab255450)
Lane 1: Wild-type A549 Control IFN-gamma (0 ng/mL, 48 h) at 20 µg
Lane 2: Wild-type A549 Treated IFN-gamma (100 ng/mL, 48 h) at 20 µg
Lanes 3 - 4: Western blot - Human CD274 (PD-L1) knockout A549 cell line (Human CD274 (PD-L1) knockout A549 cell line ab267055)
Lane 3: CD274 knockout A549 Control IFN-gamma (0 ng/mL, 48 h) at 20 µg
Lane 4: CD274 knockout A549 Treated IFN-gamma (100 ng/mL, 48 h) at 20 µg
Secondary
All lanes: Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 33 kDa
Observed band size: 45 kDa