Estimation of morphological characteristics of single particles from light scattering data in flow cytometry

Numerical methods for solving the reverse problem of light scattering for single particles and a variety of experimental results have been reviewed. In particular, the two-dimensional Mie scattering method for the estimation of sizes and refractive indices of single particles has been developed. The method of triple two-dimensional Mie scattering has been elaborated, which extends the range of particle sizes with correct solutions of the reverse light scattering problem. The flying light scattering indicatrix method, which allows the estimation of the parameters of spherical particles, is presented. The results of the use of a flow cytometer of standard design for the anaysis of milk microflora and for an AIDS immunoassay by latex agglutination are given. The design of a scanning flow cytometer is considered. The results of measuring the light scattering from polystyrene latex particles are given.Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 7, pp. 1181–1190, July, 1994.The author is grateful to A. K. Petrov and M. A. Kamkhe for supporting the work on cytometry, E. G. Saprykin for useful discussion and comments, G. Alm (Swedish Agricultural Univ.) for initiating the interest to immunological studies, and A. V. Chernyshev, T. S. Mashnin, V. I. Prots, and A. A. Doroshkin for cooperation.     

Association of calcium phosphate and fibroblast growth factor-2: a dynamic light scattering study

The size distributions of fibroblast growth factor-2 (FGF-2) in aqueous solutions with neutral pH were investigated with a dynamic light scattering technique. We found that the FGF-2 was distributed in dimer or trimer form at concentrations of 0.1-1.0 mg . mL(-1). An aggregate with a hydrodynamic radius of approximately 90 nm coexisted with this and its proportion increased with a decrease in concentration. At lower concentrations (less than 0.10 mg . mL(-1)) FGF-2 aggregates with an average radius of 80-100 nm were dominant and were stable for more than a day. These FGF-2 solutions were mixed with calcium phosphate solutions to produce a sub-micron sized compound of FGF-2 and hydroxyapatite, which could be used as a biological implant that possessed a pharmacological function for bone formation. By utilizing a transformation from amorphous calcium phosphate to hydroxyapatite, FGF-2 was effectively incorporated into polycrystals of hydroxyapatite.SEM photograph of a mixture of hydroxyapatite and FGF-2.     

Dynamic and electrophoretic light scattering of poly(dimethyldiallylammonium chloride) in salt-free solutions

Dynamic and electrophoretic light scattering were used to study the diffusion and electrophoretic mobility of poly(dimethyldiallylammonium chloride) as a function of polymer molecular weight in salt-free solutions. Two relaxation modes characterized as fast diffusion (D_f) and slow diffusion (D_s) were obtained from dynamic light scattering. Although the slow diffusion coefficient D_s strongly depends on molecular weight (M_w), the fast diffusion coefficient D_f was found to be independent of M_w over the range in the study. The fast diffusion was considered as the diffusion of a part of the polymer chain; the slow diffusion was interpreted by multichain diffusion. Electrophoretic light scattering results in the salt-free solution show that the electrophoretic mobility of the polymer is independent of M_w. © 1996 John Wiley & Sons, Inc.     

Quantitative determination of large structures by small-angle light scattering

A new small-angle light scattering camera has been developed. In contrast to conventional detection the present system is based on a recently developed two-dimensional charge-coupled-device chip made by Thomson (France). The advantage of this chip is its excellent linear response and very low dark signal even at room temperature. The best linearity was obtained by leading each pixel signal through the same amplifying system. The optical system covered a diffraction angular region from about 1° to 15° (q = 0.2–2.6 μm⁻¹). The camera was calibrated with grids and pinholes and was tested with polymer latex particles in solution and with spherulites from polymer films. Received: 06 December 1999 Accepted: 04 February 2000     

Polyaniline dispersions 7. Dynamic light scattering study of particle formation

 When aniline is oxidized in the presence of colloidal silica, composite polyaniline–silica particles of submicrometer size are obtained. Dynamic light scattering was used to monitor the course of dispersion polymerization of aniline and the formation of particles. The increase of a hydrodynamic radius of particles was observed as polyaniline had been produced. Additional increase in particle size after polymerization has also been recorded. The rate of aniline polymerization was found to increase with increasing temperature in the range 0–50 °C. Well-defined particles are formed below 30 °C while above this temperature the colloidal stability of the resulting systems is limited. The activation energy of aniline polymerization was estimated. Received: 24 February 1997 Accepted: 7 May 1997     

Surface activity—thermodynamic properties and light scattering studies for some novel aliphatic polyester surfactants

The preparation of 12 new polyester surfactants based on aliphatic amines and different ethylene oxide content is described. These surfactants were characterized by determining their molecular weights and polydispersity by gel permeation chromatography (GPC) and nitrogen content. Drop volume tensiometry (DVT) was used to measure the surface tension at 25, 35, 45 and 55°C. The surface tension isotherms were used to determine critical micelle concentration (CMC), maximum Gibb's adsorption (Γ_max), minimum area per molecule (A_min), the effectiveness of surface tension reduction (π_cmc) and the efficiency (pC₂₀). The thermodynamic parameters of micellization (ΔG_mic, ΔH_mic, ΔS_mic) and of adsorption (ΔG_ad, ΔH_ad, ΔS_ad) were calculated and the data showed that these surfactants favor micellization to adsorption. The static scattered light intensity measurements provide the calculation of the molecular weight of micelle and the aggregation number (N°), while the dynamic light scattering provide the hydrodynamic radius of micelle (R_H) and the diffusion coefficient at different surfactant concentrations. The hydrodynamic radius of micelle (R_H) at different surfactant concentrations could be used also to determine the CMC giving results that are comparable to those obtained by surface tension measurements. All the data are discussed regarding the chemical structure of the polymeric surfactants. Copyright © 2004 John Wiley & Sons, Ltd.     

Determination of proteins in human serum by combination of flow injection sampling with resonance light scattering technique

A flow injection method combined with Resonance light scattering detection was developed for the determination of protein concentration in human serum samples. This method is based on the enhanced RLS signals of protein binding with the dye acid chrome blue K. The enhanced RLS intensities at 264 nm, in an acidic aqueous solution, were proportional to the protein concentration over the range of 2.0–40.0 μg·mL⁻¹ for human serum albumin (HSA) and the limit of detection (3σ) is 85 ng·mL⁻¹. This method was successfully applied to the quantification of total proteins in human serum samples. The maximum relative standard deviation is less than 2% and the recovery is between 97 and 103% for the standard addition method. The sample throughput was 60 h⁻¹.     

Evaluation of light scattering models to characterize concentrated polymer particles embedded in a solid polymer matrix

This work deals with the application of the static light scattering (SLS) model of Vrij (VM) for the characterization of a spherical polydisperse concentrated polymer particle system. This model is the exact solution for the SLS of such mixture of particles in the Percus–Yevick approximation. The analyzed polymer particle samples are obtained by solution polymerization of isobornyl methacrylate in polyisobutylene. At the end of the polymerization, as a result of phase separation, a particle system of micrometer sized particles with a moderate distribution of sizes and a volume fraction between 5 and 10% is formed. The SLS data were also analyzed using the local monodisperse approximation (LMA), a well‐known approximation to the model of Vrij. As expected, the estimations with the VM gave better results than those performed with the LMA model for the parameters related to the shape of the particle size distribution as compared with independent determinations of these quantities obtained from scanning electron microscopy micrographs. However, the main motivation to use the more rigorous model seems to be the fact that the volume fraction of particles can be extracted from the data even when relative SLS measurements are used. © 2010 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 48: 958–963, 2010     

Evidence for the γ‐relaxation in the light scattering spectra of poly (n‐hexyl methacrylate) near the glass transition

The full range of relaxation processes present in optically pure poly‐(n‐hexyl methacrylate) (PHMA) was studied using Rayleigh–Brillouin and photon correlation spectroscopy (PCS). Brillouin shifts, linewidths, and Landau–Placzek ratios (LPR) were measured over the temperature range from ?11 to 21 °C. The Brillouin splitting and linewidth were consistent with previous studies of PHMA, but the LPR was much lower, indicating that the scattered light primarily comes from intrinsic density fluctuations. Relaxation functions of the same PHMA sample were measured using PCS over the temperature range 0.5–52.5 °C. The average relaxation times calculated from a Williams–Watts fit follow a VFT temperature dependence, with the stretching parameter β decreasing with decreasing temperature. The distribution of relaxation times reveals a merging of the α and β‐relaxations over this temperature range, and the temperature dependent width confirms that there are at least two processes with separate temperature dependences. Furthermore, there appears a process at short times in the correlation function window at low temperatures. This upturn at the fastest relaxation times is attributed to the γ‐relaxation present in higher order methacrylate polymers. The effect of the γ‐relaxation is discussed in terms of the dynamic behavior over 12 decades in time. © 2005 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 43: 1504–1519, 2005     

Unusual dispersion of the electro-optically determined electric polarizability of bentonite dispersed particles in the range 10 kHz — 1 MHz

An unusual dispersion of the electric light scattering by aqueous calcium bentonite is observed. The electro-optic effect rises with the increase of the frequency in the range 10 kHz — 1 MHz. Parallel measurements are made on suspensions of sodium bentonite and zetlizer kaolinite at different ionic strengths and particle dimensions. It is shown that the dispersion for calcium bentonite does not greatly depend on the concentration of the indifferent electrolyte (sodium chloride with concentrations 10^–4 and 5.10^–4 M/l) or on the suspension concentration. Several possibilities for explaning this unusual dispersion are considered.     

Complexation of poly(vinyl alcohol)–congo red aqueous solutions. 3. Dynamic light scattering study

Mesoscopic structures of poly(vinyl alcohol)(SINGLEBOND)Congo red (PVA(SINGLEBOND) CR) complexes in aqueous solutions were investigated in terms of dynamic light scattering (DLS) technique. The intensity-intensity time correlation function, g⁽²⁾(t), was analyzed with an equation including a single and a stretched exponential function. Two diffusion coefficients, D_f (fast) and D_s (slow) were evaluated. D_f was converted to the apparent correlation length, ξ_app, via the mode-mode coupling hypothesis. The estimated ξ_app was insensitive to the sol(SINGLEBOND)to(SINGLEBOND)gel transition but decreased with CR concentration. This change may be related to the electrostatic screening effect. On the other hand, D_s oscillates with increasing CR concentration at a specific PVA concentration range. This explains well the reentrant sol(SINGLEBOND)gel(SINGLEBOND)sol(SINGLEBOND)gel transition behavior observed in the PVA(SINGLEBOND)CR systems. D_s seems to represent the mobility of the PVA(SINGLEBOND)CR complexes, which annihilates at the gel point. © 1996 John Wiley & Sons, Inc.     

Electric response properties of a confined gas of independent particles acted upon by a direct current electric field

Exact results for linear and nonlinear electric response properties of a non-interacting ensemble of charged particles, confined within an impenetrable box and subjected to a static, homogeneous electric field, are derived and discussed. Received: 16 September 1999 / Accepted: 3 February 2000 / Published online: 5 June 2000     

Density Functional Theory calculations on the magnetic properties of the model tyrosine radical-histidine complex mimicking tyrosyl radical Y<Subscript>D</Subscript><Superscript>·</Superscript> in photosystem II

Results of Density Functional Theory (DFT) theoretical investigations, which use a model tyrosyl (Tyr) radical and tyrosyl-histidine (Tyr-His) complex to mimick the Y _D ^· radical in Photosystem II (PSII) are presented and compared to experimental results from ¹⁵N Electron-Nuclear Double Resonance spectroscopy (ENDOR) studies of the τ nitrogen coupling from His-189 in the PSII Tyr-His complex. The DFT calculations are performed using an optimized geometry of the tyrosine radical and Tyr-His complex. The conformational space of the Tyr-His tandem is explored by varying the relative geometry of the two components; relevant parameters, such as the spin distribution on the phenoxy-ring carbons of the Tyr radical and the EPR hyperfine tensors, are calculated at each geometry and compared with the available experimental data. The isotropic ¹⁵N-ENDOR signal arising from spin delocalization on the His hydrogen-bonded to the PSII tyrosine radical is analyzed in terms of the DFT obtained parameters. The calculations of the g tensor using the Gauge Independent Atomic Orbital (GIAO) approach are presented and the influence of the geometry of the Tyr-His complex on the deviation of the g-tensor elements from the free electron values is discussed.     

木质素桃红与蛋白质作用的共振光散射研究

研究了木质素桃红与蛋白质结合的共振光散射光谱. 实验结果表明, 在pH 2.56的酸性介质中, 木质素桃红与蛋白质发生静电作用产生以282.0、 346.0、 420.0 nm以及570.0 nm为特征峰的共振光散射（RLS）增强光谱. 在570.0 nm波长光激发下, 蛋白质的质量浓度与增强共振光散射强度ΔIRLS呈线性关系, 对BSA和HSA的检出限分别为39.0和22.3 ng/mL. 方法已用于尿样中总蛋白质分析.     

Experimental study of short- and long-time diffusion regimes of spherical particles in carboxymethylcellulose solutions

The behavior of latex particles in carboxymethylcellulose (CMC) solutions with different probe radii and solution concentrations is investigated. The diffusion processes of probe particles are studied by means of dynamic light scattering experiments. The existence of two diffusion regimes (a short-time regime and a long-time regime) is established for concentrations higher than the threshold value which changes for each probe dimension. Information is also provided about the relative weight of the probe population undergoing the two diffusion processes. This experimental evidence is discussed in connection with the existence of a defined volume (cage) in which short-time diffusion is the characteristic motion, while the long-time diffusion process dominates the motion over all the cages. Furthermore, there was found to be some deviation from Stokes-Einstein behavior.     

Structure formation of surfactants in concentrated sulphuric acid: a light scattering study

A common cationic surfactant,n-hexadecylammonium hydrogensulphate, dissolved in concentrated sulphuric acid, has been studied by static and dynamic light scattering. Micelle formation has been observed even in this unusual solvent. An apparent molar mass of 45 500±4.5% was found for the aggregates. A translational diffusion coefficientD ₀=5.5×10^–9 cm²/s was measured which gave a hydrodynamically effective radius ofR _h=17.7 nm. The geometric radius of gyration wasR _g=76.2 nm. The ratioR _g/R _h=4.33 is indicative for rodlike structures. Assuming a polydispersity ofL _w/L _n=2 this corresponds to a cylinder ofL _w=152 nm. An axial ratiop _w=(L _w/d)=60.4 nm was estimated which leads to a cylinder diameter of 2.53 nm. At surfactant concentrations higher than 5% (w/vol) the rod-like micelles aggregate to form more globular structures. The time correlation function, recorded by dynamic light scattering, exhibited a two-step decay which indicates a bimodal distribution of particle sizes. The fast motion coincides with that of the micelles at low concentrations while the other is slower than the fast one by three orders of magnitude and corresponds to the translational motion of large clusters.     

Quasielastic light scattering study of chemically crosslinked gelatin solutions and gels

Quasielastic light scattering measurements are reported for experiments performed on mixtures of gelatin and glutaraldehyde (GA) in the aqueous phase, where the gelatin concentration was fixed at 5 (w/v) and the GA concentration was varied from 1×10⁻⁵ to 1×10⁻³ (w/v). The dynamic structure factor, S(q,t), was deduced from the measured intensity autocorrelation function, g ₂(τ), with appropriate allowance for heterodyning detection in the gel phase. The S(q,t) data could be fitted to S(q,t)=Aexp(−D _f q ² t)+Bexp(−t/τ_c)^β, both in the sol (50 and 60 ^∘C) and gel states (25 and 40 ^∘C). The fast-mode diffusion coefficient, D _f showed almost negligible dependence on the concentration of the crosslinker GA; however, the resultant mesh size, ξ, of the crosslinked network exhibited strong temperature dependence, ξ∼(0.5−χ)^1/5exp(−A/RT) implying shrinkage of the network as the gel phase was approached. The slow-mode relaxation was characterized by the stretched exponential factor exp(−t/τ_c)^β. β was found to be independent of GA concentration but strongly dependent on the temperature as β=β₀+β₁ T+β₂ T ². The slow-mode relaxation time, τ_c, exhibited a maximum GA concentration dependence in the gel phase and at a given temperature we found τ_c(c)=τ₀+τ₁ c+τ₂ c ². Our results agree with the predictions of the Zimm model in the gel case but differ significantly for the sol state. Received: 25 May 1999 /Accepted in revised form: 27 July 1999     

Laser light scattering study of the degradation of poly(sebacic anhydride) nanoparticles

Poly(sebacic anhydride) (PSA) is biocompatible and degradable in basic media. We micronized this water‐insoluble polymer into stable polymeric nanoparticles via a microphase inversion. Such PSA nanoparticles degraded much faster than bulk PSA. The influence of the surfactant, temperature, and pH on the degradation of the PSA nanoparticles was investigated by a combination of static and dynamic laser light scattering. Under each condition, the degradation rate was nearly constant up to a 75% weight loss; that is, the degradation was close to zero‐order. The degradation rate increased with the pH and temperature. Biomedical applications of such PSA nanoparticles are suggested. © 2001 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 39: 703–708, 2001     

Interactions of norfloxacin with DNA and determination of DNA at nanogram levels based on the measurement of enhanced resonance light scattering

A novel assay of DNA at nanogram levels is presented that is based on the measurement of resonance light scattering (RLS) signals in the presence of norfloxacin. The characteristics of RLS spectra, the effective factors and the optimum conditions of the reaction have been investigated. In Britton-Robinson buffer (pH 5.87), norfloxacin has a maximum peak at 405.5 nm and the RLS intensity is greatly enhanced by trace amounts of DNA due to the interaction between norfloxacin and DNA. Mechanistic studies show that the binding of norfloxacin to DNA forms large particles, which result in the significant enhancement of RLS intensity. The enhanced intensity of RLS is proportional to the concentration of DNA in the range from 0.01–2.0 μg mL⁻¹ for yeast DNA, and from 0.02 to 2.3 μg mL⁻¹ for calf thymus DNA. The determination limit (3σ) is 0.7 ng mL⁻¹ for yeast DNA and 1.2 ng mL⁻¹ for calf thymus DNA, respectively. Synthetic samples were determined satisfactorily.     

花菁-脱氧核糖核酸的共振光散射光谱及其分析应用

研究了一种苯并噻唑阳离子花菁与脱氧核糖核酸(DNA)作用的共振光散射光谱,在pH 6.0的六次甲基四胺-HCl缓冲介质中,痕量DNA的加入使花菁在590nm的共振光散射强度显著增强。在最佳实验条件下,增强的共振光散射强度与DNA浓度具有良好的线性关系,据此建立了一种测定DNA的共振光散射光谱法。方法的线性范围为:小牛胸腺DNA(CT DNA),0～20μg/mL,鱼精子DNA(FS DNA),0～15μg/mL;检出限分别为0.005μg/mL和0.008μg/mL。该方法已用于合成样品中DNA的测定。