Studying Flow Close to an Interface by Total Internal Reflection Fluorescence Cross Correlation Spectroscopy: Quantitative Data Analysis

Total Internal Reflection Fluorescence Cross Correlation Spectroscopy (TIR-FCCS) has recently (S. Yordanov et al., Optics Express 17, 21149 (2009)) been established as an experimental method to probe hydrodynamic flows near surfaces, on length scales of tens of nanometers. Its main advantage is that fluorescence only occurs for tracer particles close to the surface, thus resulting in high sensitivity. However, the measured correlation functions only provide rather indirect information about the flow parameters of interest, such as the shear rate and the slip length. In the present paper, we show how to combine detailed and fairly realistic theoretical modeling of the phenomena by Brownian Dynamics simulations with accurate measurements of the correlation functions, in order to establish a quantitative method to retrieve the flow properties from the experiments. Firstly, Brownian Dynamics is used to sample highly accurate correlation functions for a fixed set of model parameters. Secondly, these parameters are varied systematically by means of an importance-sampling Monte Carlo procedure in order to fit the experiments. This provides the optimum parameter values together with their statistical error bars. The approach is well suited for massively parallel computers, which allows us to do the data analysis within moderate computing times. The method is applied to flow near a hydrophilic surface, where the slip length is observed to be smaller than 10nm, and, within the limitations of the experiments and the model, indistinguishable from zero.Comment: 18 pages, 12 figure

Cladoceran birth and death rates estimates

I. Birth and death rates of natural cladoceran populations cannot be measured directly. Estimates of these population parameters must be calculated using methods that make assumptions about the form of population growth. These methods generally assume that the population has a stable age distribution. 2. To assess the effect of variable age distributions, we tested six egg ratio methods for estimating birth and death rates with data from thirty-seven laboratory populations of Daphnia pulicaria. The populations were grown under constant conditions, but the initial age distributions and egg ratios of the populations varied. Actual death rates were virtually zero, so the difference between the estimated and actual death rates measured the error in both birth and death rate estimates. 3. The results demonstrate that unstable population structures may produce large errors in the birth and death rates estimated by any of these methods. Among the methods tested, Taylor and Slatkin's formula and Paloheimo's formula were most reliable for the experimental data. 4. Further analyses of three of the methods were made using computer simulations of growth of age-structured populations with initially unstable age distributions. These analyses show that the time interval between sampling strongly influences the reliability of birth and death rate estimates. At a sampling interval of 2.5 days (equal to the duration of the egg stage), Paloheimo's formula was most accurate. At longer intervals (7.5–10 days), Taylor and Slatkin's formula which includes information on population structure was most accurate

Discerning Aggregation in Homogeneous Ensembles: A General Description of Photon Counting Spectroscopy in Diffusing Systems

In order to discern aggregation in solutions, we present a quantum mechanical analog of the photon statistics from fluorescent molecules diffusing through a focused beam. A generating functional is developed to fully describe the experimental physical system as well as the statistics. Histograms of the measured time delay between photon counts are fit by an analytical solution describing the static as well as diffusing regimes. To determine empirical fitting parameters, fluorescence correlation spectroscopy is used in parallel to the photon counting. For expedient analysis, we find that the distribution's deviation from a single Poisson shows a difference between two single fluor moments or a double fluor aggregate of the same total intensities. Initial studies were performed on fixed-state aggregates limited to dimerization. However preliminary results on reactive species suggest that the method can be used to characterize any aggregating system.Comment: 30 pages, 5 figure

Attenuated total reflection infrared spectroscopy for studying adsorbates on planar model catalysts : CO adsorption on silica supported Rh nanoparticles

A sensitive method is presented for studying adsorption of gaseous species on metal surfaces in vacuum by attenuated total internal reflection Fourier transform IR spectroscopy (ATR). The method is illustrated by CO adsorption expts. on silica supported Rh nanoparticles. An exptl. setup and a procedure are described in detail to obtain a sensitivity of reflectance change of .apprx.5 * 10-5 absorbance units. Here, a silicon ATR crystal with a 50 nm layer of hydroxylated silica acts as the support for the Rh nanoparticles. These particles are easily prepd. by spincoat impregnation from a RhCl3 soln. followed by H2 redn. XPS before and after redn. shows that rhodium is reduced to Rh0 and that all chlorine is removed. At. force microscope images the distribution of the particles, which are 3-4 nm in height. When the crystal is exposed to pressures up to 1 mbar of CO, a gas which is inert to the silica support, the stretch vibration of linearly adsorbed CO on the Rh nanoparticles is detected at 2023 cm-1, while no bridged CO or geminal dicarbonyl species can be distinguished. The min. detectable coverage is estd. .apprx.0.005 CO per nm2 substrate area or .apprx.5 * 10-4 ML. [on SciFinder (R)

Proinsulin C-peptide elicits disaggregation of insulin resulting in enhanced physiological insulin effects

Using surface plasmon resonance (SPR) and electrospray mass spectrometry (ESI-MS), proinsulin C-peptide was found to influence insulin-insulin interactions. In SPR with chip-bound insulin, C-peptide mixed with analyte insulin increased the binding, while alone C-peptide did not. A control peptide with the same residues in random sequence had little effect. In ESI-MS, C-peptide lowered the presence of insulin hexamer. The data suggest that C-peptide promotes insulin disaggregation. Insulin/insulin oligomer μM dissociation constants were determined. Compatible with these findings, type 1 diabetic patients receiving insulin and C-peptide developed 66% more stimulation of glucose metabolism than when given insulin alone. A role of C-peptide in promoting insulin disaggregation may be important physiologically during exocytosis of pancreatic β-cell secretory granulae and pharmacologically at insulin injection sites. It is compatible with the normal co-release of C-peptide and insulin and may contribute to the beneficial effect of C-peptide and insulin replacement in type 1 diabetics

How does community context influence coalitions in the formation stage? a multiple case study based on the Community Coalition Action Theory

<p>Abstract</p> <p>Background</p> <p>Community coalitions are rooted in complex and dynamic community systems. Despite recognition that environmental factors affect coalition behavior, few studies have examined how community context impacts coalition formation. Using the Community Coalition Action theory as an organizing framework, the current study employs multiple case study methodology to examine how five domains of community context affect coalitions in the formation stage of coalition development. Domains are history of collaboration, geography, community demographics and economic conditions, community politics and history, and community norms and values.</p> <p>Methods</p> <p>Data were from 8 sites that participated in an evaluation of a healthy cities and communities initiative in California. Twenty-three focus groups were conducted with coalition members, and 76 semi-structured interviews were conducted with local coordinators and coalition leaders. Cross-site analyses were conducted to identify the ways contextual domains influenced selection of the lead agency, coalition membership, staffing and leadership, and coalition processes and structures.</p> <p>Results</p> <p>History of collaboration influenced all four coalition factors examined, from lead agency selection to coalition structure. Geography influenced coalition formation largely through membership and staffing, whereas the demographic and economic makeup of the community had an impact on coalition membership, staffing, and infrastructure for coalition processes. The influence of community politics, history, norms and values was most noticeable on coalition membership.</p> <p>Conclusions</p> <p>Findings contribute to an ecologic and theory-based understanding of the range of ways community context influences coalitions in their formative stage.</p

Fluorescence correlation spectroscopy of the binding of nucleotide excision repair protein XPC-hHr23B with DNA substrates

The interaction of the nucleotide excision repair (NER) protein dimeric complex XPC-hHR23B, which is implicated in the DNA damage recognition step, with three Cy3.5 labeled 90-bp double-stranded DNA substrates (unmodified, with a central unpaired region, and cholesterol modified) and a 90-mer single-strand DNA was investigated in solution by fluorescence correlation spectroscopy. Autocorrelation functions obtained in the presence of an excess of protein show larger diffusion times (τ d) than for free DNA, indicating the presence of DNA-protein bound complexes. The fraction of DNA bound (θ), as a way to describe the percentage of protein bound to DNA, was directly estimated from FCS data. A significantly stronger binding capability for the cholesterol modified substrate (78% DNA bound) than for other double-stranded DNA substrates was observed, while the lowest affinity was found for the single-stranded DNA (27%). This is in accordance with a damage recognition role of the XPC protein. The similar affinity of XPC for undamaged and 'bubble' DNA sub

Cellular mechanisms by which proinsulin C-peptide prevents insulin-induced neointima formation in human saphenous vein

AIMS/HYPOTHESIS: Endothelial cells (ECs) and smooth muscle cells (SMCs) play key roles in the development of intimal hyperplasia in saphenous vein (SV) bypass grafts. In diabetic patients, insulin administration controls hyperglycaemia but cardiovascular complications remain. Insulin is synthesised as a pro-peptide, from which C-peptide is cleaved and released into the circulation with insulin; exogenous insulin lacks C-peptide. Here we investigate modulation of human SV neointima formation and SV-EC and SV-SMC function by insulin and C-peptide. METHODS: Effects of insulin and C-peptide on neointima formation (organ cultures), EC and SMC proliferation (cell counting), EC migration (scratch wound), SMC migration (Boyden chamber) and signalling (immunoblotting) were examined. A real-time RT-PCR array identified insulin-responsive genes, and results were confirmed by real-time RT-PCR. Targeted gene silencing (siRNA) was used to assess functional relevance. RESULTS: Insulin (100 nmol/l) augmented SV neointimal thickening (70% increase, 14 days), SMC proliferation (55% increase, 7 days) and migration (150% increase, 6 h); effects were abrogated by 10 nmol/l C-peptide. C-peptide did not affect insulin-induced Akt or extracellular signal-regulated kinase signalling (15 min), but array data and gene silencing implicated sterol regulatory element binding transcription factor 1 (SREBF1). Insulin (1-100 nmol/l) did not modify EC proliferation or migration, whereas 10 nmol/l C-peptide stimulated EC proliferation by 40% (5 days). CONCLUSIONS/INTERPRETATION: Our data support a causative role for insulin in human SV neointima formation with a novel counter-regulatory effect of proinsulin C-peptide. Thus, C-peptide can limit the detrimental effects of insulin on SMC function. Co-supplementing insulin therapy with C-peptide could improve therapy in insulin-treated patients

Uptake and depuration of gold nanoparticles in Daphnia magna

This study presents a series of short-term studies (total duration 48 h) of uptake and depuration of engineered nanoparticles (ENP) in neonate Daphnia magna. Gold nanoparticles (Au NP) were used to study the influence of size, stabilizing agent and feeding on uptake and depuration kinetics and animal body burdens. 10 and 30 nm Au NP with different stabilizing agents [citrate (CIT) and mercaptoundecanoic acid (MUDA)] were tested in concentrations around 0.5 mg Au/L. Fast initial uptake was observed for all studied Au NP, with CIT stabilized Au NP showing similar rates independent of size and MUDA showing increased uptake for the smaller Au NP (MUDA 10 nm > CIT 10 nm, 30 nm > MUDA 30 nm). However, upon transfer to clean media no clear trend on depuration rates was found in terms of stabilizing agent or size. Independent of stabilizing agent, 10 nm Au NP resulted in higher residual whole-animal body burdens after 24 h depuration than 30 nm Au NP with residual body burdens about one order of magnitude higher of animals exposed to 10 nm Au NP. The presence of food (P. subcapitata) did not significantly affect the body burden after 24 h of exposure, but depuration was increased. While food addition is not necessary to ensure D. magna survival in the presented short-term test design, the influence of food on uptake and depuration kinetics is essential to consider in long term studies of ENP where food addition is necessary. This study demonstrates the feasibility of a short-term test design to assess the uptake and depuration of ENP in D. magna. The findings underlines that the assumptions behind the traditional way of quantifying bioconcentration are not fulfilled when ENPs are studied.Peer reviewed: YesNRC publication: Ye

Diffusion in Model Networks as Studied by NMR and Fluorescence Correlation Spectroscopy

We have studied the diffusion of small solvent molecules (octane) and larger hydrophobic dye probes in octane-swollen poly(dimethyl siloxane) linear-chain solutions and end-linked model networks, using pulsed-gradient nuclear magnetic resonance (NMR) and fluorescence correlation spectroscopy (FCS), respectively, focusing on diffusion in the bulk polymer up to the equilibrium degree of swelling of the networks, that is, 4.8 at most. The combination of these results allows for new conclusions on the feasibility of different theories describing probe diffusion in concentrated polymer systems. While octane diffusion shows no cross-link dependence, the larger dyes are increasingly restricted by fixed chemical meshes. The simple Fujita free-volume theory proved most feasible to describe probe diffusion in linear long-chain solutions with realistic parameters, while better fits were obtained assuming a stretched exponential dependence on concentration. Importantly, we have analyzed the cross-link specific effect on probe diffusion independently of any specific model by comparing the best-fit interpolation of the solution data with the diffusion in the networks. The most reasonable description is obtained by assuming that the cross-link effect is additive in the effective friction coefficient of the probes. The concentration dependences as well as the data compared at the equilibrium degrees of swelling indicate that swelling heterogeneities and diffusant shape have a substantial influence on small-molecule diffusion in networks.