Vimentin (VIM) is an intermediate filament protein that plays a critical role in maintaining cell integrity, supporting cellular structure, and involved in various processes like wound healing, cellular migration, and epithelial-mesenchymal transition (EMT). VIM is often used as a biomarker in studies related to cancer, fibrosis, and inflammation. Vimentin ELISA kits are used to quantitatively measure vimentin levels in biological samples such as serum, plasma, tissue lysates, and cell culture supernatants.
Overview of VIM ELISA Kits
- Purpose: To detect and quantify vimentin protein levels in biological samples, often used in research on cancer, fibrosis, and other diseases associated with cellular structure and EMT.
- Assay Principle: Typically, a sandwich ELISA. Vimentin in the sample is captured by a specific antibody, followed by detection with a secondary antibody conjugated to an enzyme (usually HRP). The resulting enzyme-substrate reaction produces a color change that is directly proportional to the amount of vimentin present.
- Sensitivity: High sensitivity, capable of detecting vimentin in the nanogram per milliliter (ng/mL) range.
- Specificity: High specificity to vimentin, minimizing cross-reactivity with other proteins.
- Sample Volume: Typically requires 50-100 µL of sample per well.
Content of a Standard VIM ELISA Kit
- Microtiter Plate: Pre-coated with an anti-vimentin antibody.
- Standards: Recombinant vimentin at various concentrations to create a standard curve.
- Detection Antibody: Biotinylated or enzyme-conjugated anti-vimentin antibody.
- Enzyme Conjugate: Usually streptavidin-HRP, which binds to the biotinylated detection antibody.
- Substrate Solution: Typically TMB (3,3',5,5'-Tetramethylbenzidine), which produces a measurable color change when reacted with HRP.
- Stop Solution: Often a sulfuric acid solution that stops the enzymatic reaction, stabilizing the color change.
Key Steps in the Assay Procedure
- Sample/Standard Addition: Samples or standards are added to wells pre-coated with an anti-vimentin antibody.
- Incubation: Plates are incubated to allow vimentin in the samples to bind to the capture antibody.
- Washing: Unbound materials are removed by washing.
- Detection Antibody Addition: The detection antibody specific to vimentin is added, binding to the captured protein.
- Washing: Excess detection antibody is removed through additional washing.
- Enzyme Conjugate Addition: Enzyme-conjugated streptavidin is added, which binds to the biotinylated detection antibody.
Applications
- Research: Used extensively in studies involving cancer progression, EMT, wound healing, and cellular stress responses.
- Diagnostics: Vimentin levels can be assessed to study tumor invasiveness, metastatic potential, and other conditions related to cellular dynamics.
Vimentin ELISA kits are vital tools in research and diagnostics, particularly in understanding the mechanisms of cancer metastasis, fibrosis, and other diseases associated with changes in cellular structure and behavior.
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