Single-Molecule Flow Experiments Using Confocal Microscopy

This study enroll companion Brownian dynamics simulations to help explain details of our experimental observations using a parameter coarse-grained model of von Willebrand. By using single-molecule flow experiments performing confocal microscopy and a microfluidic device;for shear rates up to 20,000 and present results for the shear-induced unraveling and elongation of tethered von Willebrand factor multimers.

But in some large and mysterious multimeric glycoprotein moves through the blood, balled up tightly, its reaction sites unexposed. But when significant bleeding occurs; it springs into action, initiating the clotting process. When it works properly; von Willebrand helps stop bleeding and saves lives.

But in this study global conformation changes of tethering von Willebrand can be accurately captured using a relatively simple mechanical model. However; about one to two percent of the world’s population; is affected by von Willebrand mutations; that resulting in bleeding disorders. But for those with more rare, severe forms; But very expensive treatment in the form of blood plasma replacement will be requiring.

Protein’s Behavior For Effective Drug Delivery

For a better understanding of how von Willebrand functions; could result in drugs that replace it in those who lack it; It could also lead to the development of new drugs or drug carriers that mimic the protein’s behavior for more effective drug delivery. On the other hand; if von Willebrand activates where it isn’t needed;it can trigger a stroke or heart attack.

In this study; von Willebrand multimer sample is derived from human plasma;and the molecular weight has a wide distribution as confirmed by Western blot; A sulfo-N-hydroxysuccinimide-LC-Biotinylation kit and an Alexa 488 5-sulfodichlorophenol Ester kit applying to tag von Willebrand with biotin and fluorophore molecules.Circulating von Willebrand exhibits a wide range of sizes, from dimer units to large multimers containing up to 40 to 200 monomers.

Large Variations In The Extensional Responses

Western blot results indicated that our von Willebrand sample had a wide MW distribution starting at approximately 1000 kDa; although determination of the larger multimer MWs is difficult because of the lack of standards. Shear induced dynamics of the 70 molecule ensemble by comparing normalizing elongation data; which revealed large variations in the extensional responses of tethered von Willebrand.

Methods developed in this study are useful tools for examining flow-induced macro molecule conformation; toward understanding their biological functions.This study showing that global conformation changes of tethered von Willebrand can be predicted using a simple mechanical bead spring model.