Using a single experimental approach, namely dynamic light scattering in polyacrylamide and fibrin gels, the fluctuation dynamics of polymer gels was studied for the first time in parallel at two scales. One scale is congruent with gel network macrostructure, and another is microscopic at this level. Long-period fluctuations of the network structure with characteristic times in the range ~10–70 min have been detected, during which the microdynamics of network fibers remains practically unchanged in the range 0.1–106 μs. The limits of applicability of the “frozen” inhomogeneities approximation widely used in studying gels microdynamics have been established. The approach developed forms a basis for extending the range of investigations of polymer gel dynamics.

The effect of peroxide-induced oxidation of fibrinogen on modification of its primary structure and functional properties was investigated. The oxidation sites were shown to be Met, Trp, and His residues. Using the DLS method, it was found that the oxidative modification of fibrinogen results in the change of microrheological characteristics of fibrin network. The fibrinogen oxidation diminishes its tolerance to plasmin hydrolysis and deteriorates the factor XIIIa ability to stabilize the fibrin gel.

In order to investigate quantitatively the role of lipopolysaccharides (LPS) from outer bacterial membrane at the initial state of bacterium adhesion to a host cell membrane, a model system for single cell force spectroscopy was developed and used. The system comprised of an LPS-coated microsphere placed into optical trap and a J774 macrophage being approached the microsphere to initiate their binding and then moved back to rupture the bond. An “object shadow” phenomenon was discovered, manifested as large-scale variations of the signal of photodetector registering the trapped microsphere displacement, such variations emerging long before the actual interaction between the macrophage and microsphere. The theory and the measurements technique were developed for registration of the force required for detachment of bounded microsphere from the object investigated by means of optical tweezers under the “object shadow” conditions. Characteristic spectra of binding force between J774 macrophage and microspheres functionalized with various LPS, as well as LPS plus complementary antibodies preparations were obtained at the rate of detachment force application of 3–6 pN/s. Force spectrum characteristic of Yersinia pseudotuberculosis LPS possessing O-antigen had a maximum at ~14 pN with half-width of ~23 pN. The treatment of O-antigen with complementary antibodies resulted in transformation of this spectrum into a spectrum with maximum at ~10 pN and half-width of ~14 pN, being almost identical to the spectrum of Y. pestis LPS devoid of O-antigen, with a maximum at ~9 pN and half-width of ~13 pN. A possible mechanism of force spectra formation has been proposed under assumptions of nonspecific binding of O-antigen and probable receptor-type binding of LPS core region to the macrophage surface. The elastic modulus of macrophage envelope, as estimated using analysis of displacement of the contacting microsphere as an indenter, was ≈0.17 pN/nm.

A method has been developed for the quantitative estimation of the binding force of a model microsphere with a eukaryocyte based on the optical trap in order to study the molecular mechanism of adhesion between an individual bacterium and a host cell. The substantial role of LPS O-side chains in the adhesiveness of Yersinia pseudotuberculosis 1b to J774 macrophages has been revealed with the use of a set of microspheres functionalized with lipopolysaccharide (LPS) preparations and antibodies with different specificities. The results indicate the significance of the O-antigen as a pathogenicity factor of Y. pseudotuberculosis in colonization of a macroorganism. The developed methodical approaches can be applied to the study of molecular mechanisms of the pathogenesis of pseudotuberculosis and other infectious diseases to improve antiepidemic service.

The phenomenon of stochastic low-frequency oscillations of erythrocyte cell membrane, termed usually the flicker of erythrocytes, is reviewed. The first part describes the theoretical models of erythrocyte flickering and the registration techniques. The relations are given and analyzed which connect the shape of both the frequency and the spatial spectra of stochastic membrane oscillations with geometrical and mechanical parameters of the erythrocyte and with the ambient physical characteristics. The existing concepts of excitation mechanisms of the membrane flickering are presented.

The phenomenon of stochastic low-frequency oscillations of erythrocyte cell membrane, termed usually the flicker of erythrocytes, is reviewed. The first part [Biol. Membrany (Rus.), 2009, vol. 26, no. 5, pp. 352–369] describes theoretical models of erythrocyte flickering and the registration techniques. In the second part presented below the main experimental results are reviewed, the problem of identification of acting mechanisms of flicker excitation is analyzed, and flicker interrelations are considered with the membrane functioning as well as with the dynamics of proteins embedded in the membrane. The possibilities and the prospects of medical diagnostics applications of flicker of erythrocytes are discussed briefly.

The phenomenon of stochastic low-frequency oscillations of erythrocyte cell membrane, termed usually the flicker of erythrocytes, is reviewed. In the first part [Kononenko V.L. // Biol. membrany. 2009. V. 26. No, 5], theoretical models of erythrocyte flickering as well as registration techniques were described. The relations have been given and analyzed which connected the shape of both the frequency and the spatial spectra of stochastic membrane oscillations with the geometrical and mechanical parameters of erythrocyte and with the ambient physical characteristics. The existing concepts of excitation mechanisms of the membrane flickering were also presented. In the second part of the review given here, the main experimental data are described. The problems of identification of the acting excitation mechanisms of the membrane flickering are considered further and interrelations between the flicker and the membrane functioning as well as the dynamics of intramembrane proteins are analyzed. The possibilities and the prospects of the medical diagnostics applications of the erythrocyte flicker are discussed briefly.

An approximate theory of the erythrocyte motion due to a pressure difference in a long cylinder channel of finite length filled with a fluid is developed. The theory is based upon combined solution of equations of mechanical equilibrium of erythrocyte membrane together with fluid dynamics equations. An a priori choice of a sufficiently simple and realistic class of erythrocyte's shapes in the narrow channel with fixed diameter is used. The shape is described by three parameters - surface area and volume of erythrocyte, and the wall layer thickness. Analytical expressions are obtained for erythrocyte velocity in the channel and for the wall layer thickness as a function of geometrical and mechanical parameters of erythrocyte, as well as channel dimensions and filtration pressure. The results are a part of a developed theory of erythrocyte passage through porous filters.

An approximate non-stationary theory of erythrocyte passage (filtration) through a long narrow channel is developed. The theory is based upon solution of equations of mechanical equilibrium of erythrocyte membrane together with fluid dynamics equations for fluid in a channel and for intracellular hemoglobin solution. To describe the shape of erythrocyte in the process of filtration, an a priory choice of the geometrical class of shapes is used. Shape parameters are determined from force balance equations taking into account conservation of cell volume and surface area. A system of constitutive differential equations of filtration is obtained. The system incorporates elasticity modulii, viscosity coefficients, and geometrical parameters of erythrocyte, radius and length of a channel, filtration pressure P-f, as well as osmolarity u, temperature, and viscosity eta(b) of the surrounding fluid. Apart from numerical solutions, an approximate formula for filtration duration Delta t(er) is obtained, which depends on the parameters listed above and is convenient for qualitative analysis of the filtration process. A numerical simulation of the influence of the erythrocyte mechanical moduli on the shape of the basic filtration curves, namely, osmotic Delta t(er)(u), baric Delta t(er)(P-f), and viscosity Delta t(er)(eta(b)), is carried out.

The osmotic dependences of erythrocytes passage time through the nickel mesh with 3.1 mum diameter. 11 mum length pores are measured for healthy donors at temperatures 25, 37. 40degreesC for the medium osmolarity range 210-520 mosM. It is shown that the increase of temperature leads to a monotonic shift of the osmotic cut-off boundary of filtration curve to lower osmolarity values, as well as to the general decrease of filtration times. An approximate non-stationary theory of erythrocyte filtration through a long narrow channel is developed. The theory is based on the combined treatment of the equations of mechanical equilibrium of the cell membrane together with hydrodynamics equations for the fluid in a channel and for hemoglobin solution inside the cell. An approximate analytical solution is obtained, which describes the dependence of the transit time on the medium osmolarity, as well as on mechanical, geometric, and physical parameters of the cell, channel. and measuring conditions. Generalization of the Ross-Minton formula for the concentration dependence of viscosity of hemoglobin solution is suggested to describe the temperature effect. Analysis of measurements results using the theory developed showed that the temperature shift of hypotonic cut-off of erythrocytes filterability can be accounted for by the temperature increase of the cell surface area under practically unchanged cell volume. No sudden temperature transitions of viscosity of hemoglobin solution inside the cell, similar to those reported in literature for the similar conditions in vitro, were detected under conditions of the measurements.

The possibility is shown to monitor the transient ionic state and volume of erythrocyte suspended in low ionic strength solution (LISS) by registration of the dielectro-deformation (DD) of a cell. The adequate theoretical basis is developed. Possible registration modes are considered analytically and numerically using the theory of dielectro-deformation of erythrocytes developed previously.

The theory is developed of the stationary and transient deformations of erythrocytes by spatially homogeneous high-frequency (HF) electric field with a stationary or varied amplitude. The cell shape is approximated by a three-axes ellipsoid with time-dependent semi axes and constant volume and surface area. Both bending and shear membrane strains and stresses are taken into account. The theory can be applied to the flaccid lipid vesicles also. The dynamics of erythrocytes deformation due to a step-wise and harmonic variation of field amplitude is studied analytically and numerically. The shape of typically registered deformational curves is interrelated with mechanical, electric, and geometrical parameters of erythrocyte. The existing experimental data set both on stationary and transient deformations of erythrocytes by HF field is described quantitatively using the theory developed. Some important electric, deformational and viscous parameters of human erythrocytes placed in low ionic strength medium are determined from such description.

Stochastic bending oscillations of cell membrane and deformations of erythrocytes by high frequency electric field are very promising phenomena for medical and environmental diagnostic applications. Experimental observations showed high sensitivity of these phenomena to variations of erythrocytes mechanical properties caused by physico-chemical factors and by pathological changes in the cell. However, for unambiguous interpretation of the measured dependencies, and for adequate relation of their changes to the alterations of particular mechanical parameters of erythrocytes, a comprehensive theoretical basis is required. This work present and discusses the fundamentals of the flicker spectroscopy and dielectro- deformational spectroscopy of erythrocytes in a view of possible diagnostics applications.

Using the theoretical analysis within the framework of the proposed ellipsoidal shear electromechanical model of erythrocyte, the main mechanisms and relationships have been established and studied for the deformations of erythrocytes caused by a spatially homogeneous high-frequency electric field. The main types of the stress-strain curves characteristic of stationary and dynamic deformations caused by the rectangular-pulse and harmonic modulations of the field amplitude have been calculated. The relationship has been established between the parameters of essentially nonlinear stress-strain curves and mechanical, electric, and geometric parameters of erythrocyte. The impossibility of unlimited elongation of erythrocyte by the field, due to the conservation of the cell volume and surface area, has been shown, and the dependence of the maximum possible elongation of the cell on its volume has been calculated. It has been shown that the relaxation time of dynamic deformations of erythrocyte in the presence of an electric field considerably differs from that characteristic of the membrane material and sharply decreases with the increase of the initial elongation of the cell.

The theory of erythrocyte deformation by high-frequency (HF) electric field, based on ellipsoidal approximation for cell shape, is developed. Various possible deformational regimes of cell elongation by the field are analyzed, depending on the relation between the bending and the shear deformations of cell membrane and the magnitudes of corresponding mechanical stresses arising. The role of mechanical, electrical, and geometrical parameters of erythrocyte in formation of various parts of deformational curves (cell length-field strength) is elucidated, including the initial part and the geometrical saturation region. The measurements of the stationary elongation curves of erythrocytes by Krueger and Thom [Biophys. J. 73 (1997) 2653] are analyzed quantitatively. The shape of the measured curves is described exactly by the theory for the whole range of field strengths and temperatures used. The temperature-dependent values of erythrocyte volume and cytoplasmic conductivity are obtained from this description in the range (- 15-25 degrees C).

The spectra of natural oscillations of human erythrocyte cell membranes were studied experimentally and theoretically. The measurements were carried out at room temperature for both single normal cells and erythrocyte rouleaux in a range of 0.03-500 Hz. The spectra were measured at a resolution better than 1% using two techniques: registration of spontaneous membrane oscillations induced by thermal agitation in the surrounding medium and registration of the amplitude-frequency characteristics of the forced oscillations of erythrocyte elongation induced by a high-frequency electric field with the amplitude harmonic modulation. The spectra measured by both techniques had no resonance frequencies and decreased monotonically with the frequency increase. These results are confirmed by the theory developed for the extracellular excitation mechanisms of membrane oscillations. The spectra of active oscillatory biomechanical processes were measured for comparison. These processes are ciliary beating of human bronchial epithelium and ciliary beating and artificial periodic contractions of the cytoplasm of the ciliate Spirostomum ambiguum. The quality of the resonance lines of the order of 10-20 registered may serve as estimates for the line width in search of the resonance oscillations in erythrocytes induced by active cell processes.

Optical registration technique is developed and frequency spectra are measured for dynamic shape fluctuations (flicker) in erythrocytes. Flicker spectra are studied in the range 0.03-500 Hz, using dynamic microphotometering of single cells in two optical modes. phase contrast regime, and reflection-mode laser probing. The registered spectra are similar to those typical of 1/f noise, with an essential difference: their slope in log-log scale varies with frequency, from -(0.8 divided by 1.2) below 10 Hz to -(1.6 divided by 2.4) above 50 Hz. The spectra measured with backward laser light scattering go systematically higher then those measured in the phase contrast regime. The theory of erythrocyte flicker is developed, based on the eigenmodes analysis of the bending oscillations of the cell envelope, and on the optical problem solution for the contribution from these oscillations to the registered signal. The theory describes quantitatively the spectral curves, and relates their parameters to the main mechanical and shape parameters of erythrocytes: bending modulus of cell membrane, viscosity coefficient of hemoglobin solution, and thickness-diameter ratio of a cell. The difference in the spectra obtained by coherent and noncoherent probing of erythrocytes is explained by specificity of optical mechanisms of the registered signal formation. No firm evidence is obtained for the contribution from the active processes in erythrocytes to the flicker.