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A new TIRF single molecule calibration

Thursday, November 07, 2013 — Poster Session II

12:00 p.m. – 2:00 p.m.

FAES Academic Center (Upper-Level Terrace)




  • A. Popescu-Hategan


A new total internal reflection fluorescence (TIRF) molecular calibration based on single molecule fluorescence is presented. Used to study myosin polymerization, it determined the number of molecules in each single filament formed in the presence or absence of a copolymerizing protein. Exchange and depolymerization were observed live in single filaments and their rates measured: first experimental evidence that exchange takes place by monomer addition as determined by single molecule calibration. Live observation of fibrin polymerization in TIRF was translated by this calibration in molecular information on the growth kinetics of fibers. Fibrin fibers thickened in time to thousands of molecules across, whereas some fibers reached early a stationary state. Diameters of fibers were calculated from this data and their evolution observed during growth to go up to hundreds of nm. This new approach in protein polymerization bridges the gap between molecular and assembled structure by determining macroscopic features of the polymer starting from single molecule observation thus providing information important for understanding the early events of polymerization.

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