Anti-Vinculin Monoclonal Antibody for WB, IF-P, ELISA - P18206

Core Product Specifications and Parameters

Biological Background: Vinculin Function and Localization

• Vinculin is a highly conserved actin-binding protein that links integrins and cadherins to the actin cytoskeleton, playing key roles in both cell-matrix and cell-cell adhesion.
• A muscle-specific isoform, known as metavinculin (MV), arises from alternative splicing and contains an additional exon; it is predominantly expressed in cardiac and skeletal muscle.
• Vinculin localizes to cell-matrix focal adhesions, cell-cell adherens junctions, and the perinuclear region, as well as to specialized structures such as podosomes and sarcolemma.
• The protein undergoes multiple post-translational modifications, including phosphorylation, acetylation, and palmitoylation, which regulate its activation, protein interactions, and membrane targeting.
• Vinculin acts as a mechanosensor; tension-induced conformational changes expose cryptic binding sites for talin and F-actin, thereby strengthening adhesion complexes.
• As a key component of the cadherin-catenin complex, vinculin modulates E-cadherin surface expression and potentiates mechanosensing at adherens junctions, impacting tissue morphogenesis and homeostasis.
• Consistent with its essential roles in adhesion and mechanotransduction, mutations in VCL are associated with dilated cardiomyopathy (CMD1W) and hypertrophic cardiomyopathy (CMH15).
• Additionally, vinculin participates in cell morphology, migration, and invasion, as evidenced by its recruitment to podosomes and its influence on lamellipodial dynamics.

Experimental Guidance and Technical Tips

• The immunogen is a synthetic peptide corresponding to the C-terminal region (aa800-900) of human Vinculin; this region is conserved across human, mouse, and rat, explaining the reported cross-reactivity.
• For Western blotting, vinculin migrates at an apparent molecular weight of ~124 kDa; use appropriate gel percentages (e.g., 8% SDS-PAGE) and verify transfer efficiency using total protein staining.
• In immunofluorescence (IF-P), consider fixation and permeabilization conditions that preserve the architecture of focal adhesions and adherens junctions (e.g., 4% PFA followed by 0.1% Triton X-100). Co-staining with paxillin or F-actin markers can confirm localization.
• When performing ELISA, titrate the antibody against the immunogen peptide and native protein to determine optimal concentrations; cross-adsorption may be evaluated if high background is observed in complex matrices.
• As vinculin is a ubiquitous protein, including appropriate loading controls (e.g., GAPDH, β-actin) is essential for quantitative analyses.
• For studies involving muscle tissues, note that the antibody may recognize both vinculin and metavinculin isoforms; the latter runs slightly higher at approximately 150 kDa.

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