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In vivo vascular leakage assay
1Institute of General Pathology, University of Milan, Milan, Italy.
Methods in Molecular Medicine
|April 6, 2004
Summary
This study details a method to measure increased vascular permeability using animal models. By perfusing organs with dye and albumin, researchers can quantify leakage, aiding in the study of inflammatory mediators like tumor necrosis factor alpha (TNFalpha).
Area of Science:
- Physiology
- Pharmacology
- Biomedical Engineering
Background:
- Vascular permeability is a critical physiological parameter.
- Increased vascular permeability is associated with inflammation and disease.
- Tumor necrosis factor alpha (TNFalpha) is a key mediator of inflammation.
Purpose of the Study:
- To describe a method for quantifying in vivo vascular permeability.
- To assess the impact of inflammatory mediators like TNFalpha on vascular leakage.
- To establish a reproducible assay for measuring endothelial barrier function.
Main Methods:
- In vivo perfusion of mouse liver or kidney via portal vein or renal artery.
- Intravascular injection of blue dye (trypan blue or Evans blue) complexed with albumin.
- Spectrophotometric analysis (540 nm) of homogenized organ supernatants after saline washing to quantify extravasated dye.
Main Results:
- Increased optical density in supernatants correlates with dye extravasation.
- Dye retention in subendothelial spaces indicates increased endothelial permeability.
- The method allows for quantitative measurement of vascular leakage.
Conclusions:
- This perfusion-based method provides a reliable means to measure TNFalpha-induced vascular permeability in vivo.
- The technique is applicable to studying other mediators affecting endothelial barrier function.
- Quantification of dye retention serves as a direct indicator of vascular leakage.