Surface ionization of metastable calcium and ytterbium atoms

This dissertation presents an experiment demonstrating the surface ionization of metastable atoms on a tungsten surface. Surface ionization refers to the thermionic ionization of adsorbates with low ionization potentials on hot, high-work function metal surfaces. The efficiency of surface ionization is exponentially dependent on the difference between the atom ionization energy and the surface work function. Since the work functions of metals are, at most, ~ 6 eV, surface ionization is limited to elements with low ionization energies and is frequency exploited for detecting alkali and alkaline earth metal beams. We use an electric discharge to excite a fraction of atoms in separate calcium and ytterbium beams to metastable states. In doing so, we effectively lower their ionization energy by about 2 eV. On a polycrystalline tungsten surface, with a work function measured to be 5.18 eV, surface ionization is theoretically more likely for metastable calcium and ytterbium atoms than for their ground state counterparts. However, the elementary theory of surface ionization does not take into account various de-excitation or neutralization processes that may preclude the formation of ions. We report findings that metastable calcium and ytterbium atoms undergo surface ionization at significantly higher rates than corresponding ground state atoms.Physic

Physical Determinants of Fibrinolysis in Single Fibrin Fibers

Fibrin fibers form the structural backbone of blood clots; fibrinolysis is the process in which plasmin digests fibrin fibers, effectively regulating the size and duration of a clot. To understand blood clot dissolution, the influence of clot structure and fiber properties must be separated from the effects of enzyme kinetics and perfusion rates into clots. Using an inverted optical microscope and fluorescently-labeled fibers suspended between micropatterned ridges, we have directly measured the lysis of individual fibrin fibers. We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected. Thrombin and plasmin dose-response experiments showed that the elongation behavior was independent of plasmin concentration, but was instead dependent on the concentration of thrombin used during fiber polymerization, which correlated inversely with fiber diameter. Thinner fibers were more likely to lyse, while fibers greater than 200 ± 30 nm in diameter were more likely to elongate. Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain. Using polymer physics- and continuum mechanics-based mathematical models, we show that fibers polymerize in a strained state and that thicker fibers lose their prestrain more rapidly than thinner fibers during lysis, which may explain why thick fibers elongate and thin fibers lyse. These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities

Submillisecond Elastic Recoil Reveals Molecular Origins of Fibrin Fiber Mechanics

ABSTRACT Fibrin fibers form the structural scaffold of blood clots. Thus, their mechanical properties are of central importance to understanding hemostasis and thrombotic disease. Recent studies have revealed that fibrin fibers are elastomeric despite their high degree of molecular ordering. These results have inspired a variety of molecular models for fibrin’s elasticity, ranging from reversible protein unfolding to rubber-like elasticity. An important property that has not been explored is the timescale of elastic recoil, a parameter that is critical for fibrin’s mechanical function and places a temporal constraint on molecular models of fiber elasticity. Using high-frame-rate imaging and atomic force microscopy-based nanomanipulation, we measured the recoil dynamics of individual fibrin fibers and found that the recoil was orders of magnitude faster than anticipated from models involving protein refolding. We also performed steered discrete molecular-dynamics simulations to investigate the molecular origins of the observed recoil. Our results point to the unstructured aC regions of the otherwise structured fibrin molecule as being responsible for the elastic recoil of the fibers

Structure of the Fibrinogen Molecule.

<p><b>A</b> structure of the fibrinogen molecule (crystal structure 3GHG) with αC domains built in using homology modeling and Discrete Molecular Dynamics Simulations [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116350#pone.0116350.ref024" target="_blank">24</a>]. Fibrinogen spans 45 nm and consists of two outer D regions, each connected by a coiled-coil segment to the central E region. Thrombin cleaves fibrinopeptides A and B from the Aα-chains and Bβ-chains, respectively, producing insoluble fibrin monomers that form fibrin networks. <b>B</b> A cartoon representation of the molecule, with the αC region highlighted in green. <b>C</b> A cartoon model of a fibrin fiber emphasizing the interaction of the αC polymer network within the fiber. </p

Modeling the lysis of a Fiber Consisting of an Inner Core and Outer Shell.

<p><b>A</b> Left: cross-section of a fiber with an outer shell (A) and inner core (B). Right: a portrayal of the relative lengths of the shell, the core, the free fiber (equilibrium fiber length), and SS-suspended fiber. <b>B</b> The fiber free length, normalized to the length between the structured surfaces, for thin (blue, 40 nm radius), medium (red, 80 nm radius), and thick (green, 120 nm radius) fibers as the shell is lysed. The black line signifies the SS length (<i>L_SS</i>) and the black dot is the free length of the fiber after 100% lysis of the shell (i.e., the core length). Note that there is no radial dependence on the free length for fibers of the constant ratio model. For this plot <i>L_oA</i> = 18 μm, <i>S</i> = 1.2. For the constant core, the core thickness was 15 nm. </p

Diameter Dependence of Fibrinolysis.

<p><b>A</b> Over two hundred single fibers were imaged on the SEM. Fiber diameters were measured at the thinnest point. Data were collected from fibers polymerized at three distinct thrombin concentrations: 0.11 U/mL (84 samples), 1.1 U/mL (64 samples), and 11 U/mL (73 samples), (p < 0.02 for all cases). <b>B</b> Percentage of fibers that lysed or elongated during the first thirty minutes of exposure to 3.3 U/mL of plasmin. Fibrin fibers polymerized by higher concentrations of thrombin lysed more frequently than those polymerized by lower thrombin concentrations. <b>C–E</b> The data in the histograms were segregated according to the percentage of fibers that lysed or elongated. The percentage of fibers with thicknesses below the <i>d</i>₀ diameters (indicated by a white dotted line) parallels the percentage of fibers that lysed in the corresponding bar graphs. The threshold diameter (<i>d₀</i>) was 200 nm ± 30 nm.</p

Effect of Plasmin concentration on Lysis and Elongation.

<p><b>A</b> The average time measured for single fibers to lyse after exposure to plasmin. Fibers exhibiting elongation were not counted in this analysis. <b>B</b> Percentage of fibers that exhibited lysis, elongation, and no change within thirty minutes after exposure to plasmin. Note that the ratio of the number of fibers that elongated to that of fibers that lysed remained constant across the range of plasmin concentrations. ‘n’ indicates the total number of fibers (lysed, elongated, no change) observed per plasmin dose. </p

Images of elongated and lysed fibrin fibers.

<p>Epi-fluorescence microscopy of fibrin fibers suspended across structured surfaces (bar = 20 μm). Samples were labeled with 20 nm red fluorescent beads and polymerized with 0.11–11 U/mL of human thrombin. Images of fibers before (A, C, E, G, I, K, M) and 5–15 minutes after (B, D, F, H, J, L, N) addition of plasmin. Samples 1–2 were treated with 20 μL of 1.0 U/mL of plasmin and display a lysed fiber (B) or a lysed and elongated fiber in the same field of view (D). Samples 3–7 were treated with 20 μL of plasmin ranging from 0.6 U/mL–6.0 U/mL and show elongated fibers, exhibiting extensions up to 10%. </p