Influence of chain charge and complexation on the overlap and entanglements formation in poly(acrylic acid) salt-containing aqueous solutions

Dilute-semidilute regime crossover in aqueous solutions of partly neutralized poly(acrylic acid) and of its complex with tetradecyltrimethylammonium bromide was studied by light scattering and viscometry methods. The chain charge growth causes the decrease of overlap concentration (c*) and the increase of the entanglements formation concentration (ce), hence, the semidilute unentangled regime of solution expands. Complexation of the polyelectrolyte with an oppositely charged surfactant leads to c* increase and to ce decrease. It is shown that in semidilute entangled solutions the surfactant acts as an effective structuring agent because of the binding of polyelectrolyte chains via surfactant micelles.     

线性聚丙烯酰胺凝胶毛细管电泳的迁移特性

使用线性聚丙烯酰胺作为筛分介质,对片段长度为80～584bp的标准DNA样品进行毛细管电泳,利用激光诱导荧光方法检测信号,荧光染料为溴化乙啶。改变电场强度100～375V/cm,得到的迁移率曲线与电场强度和DNA片段长度成复杂的函数关系,已有的经典理论模型:Ogston模型、Reptation无拉伸模型和Reptation拉伸模型都不能正确地描述实验观察到的迁移率随电场强度和DNA片段长度的变化情况。因此,提出一种修正的Ogston筛分理论,假定迁移的DNA分子在电场强度方向延展拉伸,如同小分子穿过凝胶筛孔。在该修正模型中,DNA的迁移率仅依赖于电场强度、筛分介质浓度和片段长度,很好地解释了实验现象。     

The microrheology of polystyrene sulfonate combs in aqueous solution

Video particle tracking (VPT) and diffusing wave spectroscopy were used to characterize the microrheology of polystyrene sulfonate combs in aqueous solutions. At low frequencies VPT demonstrated predominantly viscous behavior. The manner in which the viscosity scaled as a function of monomer concentration was a sensitive function of the comb architecture. Densely branched combs or combs with long side chains demonstrated entangled polyelectrolyte scaling above the overlap concentration, whereas sparsely branched combs had unentangled polyelectrolyte scaling. A dynamic scaling model was developed for the viscosity of unentangled semidilute solutions of comb polyelectrolytes. Diffusing wave spectroscopy demonstrated Rouse modes (G' approximately G" approximately omega12) for the high-frequency dynamics of the semidilute comb solutions. The form of the high-frequency viscoelasticity was independent of the chain architecture and the modulus scaled as expected for linear flexible polyelectrolytes.     

Mathematical model for DNA separation by capillary electrophoresis in entangled polymer solutions

A mathematical model of DNA separation by capillary electrophoresis in entangled polymer solution is presented. The mechanism is modeled as a DNA molecule moving through transient pores formed in polymer solutions and colliding with blobs of polymer molecules encountered during migration. By taking account of the average retardation time (t(c)) of DNA-blob collision and calculating the total collision number (N(c)), a quantitative mathematical equation was reported, leading to predictions for the DNA mobility as a function of the experimental conditions like the size of DNA, the polymer concentration and the electric field strength. For DNA fragments in frequent size range, the initial experimental data agree well with the model. The DNA shape function (f(E)) was suggested and then discussed by the experimental data. The relationship between f(E) and electric field strength E was empirically estimated. Then, the average retardation time t(c) was obtained as about (2 approximately 3)x10(-6)s in linear polyacrylamide (LPA) and hydroxyethylcellulose (HEC) solution.     

Effect of matrix chain length on the electrophoretic mobility of large linear and branched DNA in polymer solutions

We study the effect of matrix chain molecular weight Mw and concentration c on the electrophoretic mobility micro of large linear and star-like, branched DNA in polymer solutions. Polyethylene oxide (PEO) with narrow molecular weight distributions form the main focus of this study. For PEO concentrations ranging from one half the overlap concentration, c*, to 3c*, the effective drag coefficient, zeta is identical with (mu0/mu) - 1, satisfies the following approximate scaling relationship, zeta approximately cMw(0.7). Here, mu0 is the electrophoretic mobility in free solution. While the concentration dependence is consistent with predictions from the transient entanglement coupling (TEC) model, the molecular weight dependence is significantly weaker. Although a similar dependence of mobility on Mw can be predicted when nonentangling collisions are the dominant source of drag, a model based on these collisions alone cannot reproduce the experimental observations. We also find that the architecture of large DNA does not affect either the concentration dependence or molecular weight dependence of the electrophoretic mobility.     

Linear viscoelasticity of poly(acrylonitrile-co-itaconic acid)/1-butyl-3-methylimidazolium chloride extended from dilute to concentrated solutions

The solution rheology of poly(acrylonitrile-co-itaconic acid) (poly(AN-co-IA)) in 1-butyl-3-methylimidazolium chloride ([BMIM]Cl) spanning dilute, semidilute unentangled and entangled regimes were investigated. The exponents in the specific viscosity η_sp ～ overlap parameter c[η] power law were 1, 2 and 4.7 for dilute, semidilute unentangled and entangled regimes, respectively, which were found to be consistent with the scaling prediction for neutral linear polymers in θ-solvent. For dilute solutions (lower than 0.9 wt.%), the linear viscoelastic responses were observed to be in good agreement with the Zimm model (Flory exponent ν = 0.5). While for semidilute unentangled solutions (between 0.9 and 8 wt.%), results obtained had been found to be consistent with Rouse model. Considering Flory exponent ν = 0.5 and the concentration dependences of the specific viscosity and relaxation time, it had been evaluated that poly(AN-co-IA) in [BMIM]Cl behaves as a neutral polymer in θ-solvent. It had also been suggested that according to the unusual deviation of Cox-Merz rule, poly(AN-co-IA)/[BMIM]Cl solutions are typical neutral polymeric liquids for the concentrated solutions but have shown a more complicated behavior at high deformation rates.     

Sparsely cross-linked "nanogels" for microchannel DNA sequencing

We have developed sparsely cross-linked "nanogels", sub-colloidal polymer structures composed of covalently linked, linear polyacrylamide chains, as novel DNA sequencing matrices for capillary electrophoresis. The presence of covalent cross-links affords nanogel matrices with enhanced network stability relative to standard, linear polyacrylamide (LPA), improving the separation of large DNA fragments. Nanogels were synthesized via inverse emulsion (water-in-oil) copolymerization of acrylamide and N,N-methylenebisacrylamide (Bis). In order to retain the fluidity necessary in a replaceable polymer matrix for capillary array electrophoresis (CAE), a low percentage of the Bis cross-linker (< 10(-4) mol%) was used. Nanogels were characterized by multiangle laser light scattering and rheometry, and were tested for DNA sequencing by CAE with four-color laser-induced fluorescence (LIF) detection. The properties and performance of nanogel matrices were compared to those of a commercially available LPA network, which was matched for both weight-average molar mass (Mw) and extent of interchain entanglements (c/c*). Nanogels presented in this work have an average radius of gyration of 226 nm and a weight-average molar mass of 8.8 x 10(6) g/mol. At concentrations above the overlap threshold, nanogels form a clear, viscous solution, similar to the LPA matrix (Mw approximately 8.9 x 10(6) g/mol). The two matrices have similar flow and viscosity characteristics. However, because of the physical network stability provided by the internally cross-linked structure of the nanogels, a substantially longer read length ( approximately 63 bases, a 10.4% improvement) is obtained with the nanogel matrix at 98.5% accuracy of base-calling. The nanogel network provides higher-selectivity separation of ssDNA sequencing fragments longer than 375 bases. Moreover, nanogel matrices require 30% less polymer per unit volume than LPA. This is the first report of a sequencing matrix that provides better performance than LPA, in a side-by-side comparison of polymer matrices matched for Mw and extent of interchain entanglements.     

Capillary electrophoresis of RNA in dilute and semidilute polymer solutions

We report separations of RNA molecules (281-6583 nucleotides) by capillary electrophoresis in dilute and semidilute solutions of aqueous hydroxyethylcellulose (HEC) ether in varying buffers. RNA mobility and peak band widths are examined under both nondenaturing and also denaturing conditions. From studies of sieving polymer concentration and chain length, it is found that good separations can be obtained in semidilute solutions as well as in dilute solutions. The dependence of RNA mobility on its chain length is consistent with separation by a similar to transient entanglement mechanism in dilute solutions. In semidilute entangled solutions the separation proceeds by segmental motion.     

Comparison of RNA, single-stranded DNA and double-stranded DNA behavior during capillary electrophoresis in semidilute polymer solutions

We present a study of the separation of RNA, single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) in semidilute linear hydroxyethylcellulose (HEC) solution. Our results strive to provide a better understanding of the mechanisms of nucleic acid migration during electrophoresis in polymer solutions under native and denaturing conditions. From a study of the dependence of mobility on chain length and applied electric field, we found that RNA and ssDNA show better separation and higher resolution over a larger range of sizes compared to dsDNA. In addition, RNA reptation without orientation extends to longer chain lengths in comparison to ssDNA, possibly as a result of different type of short-lived secondary structure formations. Such a comparative study between nucleic acid capillary electrophoresis helps to optimize RNA separation and provides better understanding of RNA migration mechanisms in semidilute polymer solutions under denaturing conditions.     

Strain Hardening and Strain Softening of Reversibly Cross-linked Supramolecular Polymer Networks

The large amplitude oscillatory shear behavior of metallo-supramolecular polymer networks formed by adding bis-Pd(II) cross-linkers to poly(4-vinylpyridine) (PVP) in dimethyl sulfoxide (DMSO) solution is reported. The influence of scanning frequency, dissociation rate of cross-linkers, concentration of cross-linkers, and concentration of PVP solution on the large amplitude oscillatory shear behavior is explored. In semidilute unentangled PVP solutions, above a critical scanning frequency, strain hardening of both storage moduli and loss moduli is observed. In the semidilute entangled regime of PVP solution, however, strain softening is observed for samples with faster cross-linkers (k(d) ～ 1450 s(-1)), whereas strain hardening is observed for samples with slower cross-linkers (k(d) ～ 17 s(-1)). The mechanism of strain hardening is attributed primarily to a strain-induced increase in the number of elastically active chains, with possible contributions from non-Gaussian stretching of polymer chains at strains approaching network fracture. The divergent strain softening of samples with faster cross-linkers in semidilute entangled PVP solutions, relative to the strain hardening of samples with slower cross-linkers, is consistent with observed shear thinning/shear thickening behavior reported previously and is attributed to the fact that the average time that a cross-linker remains detached is too short to permit the local relaxation of polymer chain segments that is necessary for a net conversion of elastically inactive to elastically active cross-linkers. These and other observations paint a picture in which strain softening and shear thinning arise from the same set of molecular mechanisms, conceptually uniting the two nonlinear responses for this system.     

Investigation on the interaction between C16TAB and NaCMC in semidilute aqueous solution based on rheological measurement

The rheological behavior of unentangled and entangled semidilute solution of anionic polyelectrolyte sodium carboxymethyl cellulose (NaCMC) containing cationic surfactant cetyltrimethylammonium bromide (C₁₆TAB) was investigated. The results reveal that the rheological properties of these semidilute NaCMC solutions depend on the amount of C₁₆TAB added. In the unentangled semidilute NaCMC solution (0.5 g/L), the viscosity decreases with the increase of C₁₆TAB amount in the low surfactant concentration region (below the critical aggregation concentration, CAC). However, in high surfactant concentrations (above CAC), the viscosity decreases sharply with the increase in C₁₆TAB amount. It is found that viscosity change of NaCMC solution could be described using Colby’s model when surfactant concentrations are between CAC and saturated concentration (C_s), suggesting that no inter-polymer interaction exists between C₁₆TAB and NaCMC in the unentangled semidilute solution. However, for the entangled semidilute NaCMC solution (5 g/L), the addition of C₁₆TAB leads to an increase in viscosity. Meanwhile, the solution exhibits an enhanced shear thinning behavior due to adding more C₁₆TAB than 1 mM. The viscosity increase is ascribed to the physical cross linking of surfactant micelles with NaCMC chains. Furthermore, it is suggested that the enhanced shear thinning behavior results from weak interaction between NaCMC chains and C₁₆TAB micelles.     

Viscoelasticity and rheology in the regimes from dilute to concentrated in cellulose 1-ethyl-3-methylimidazolium acetate solutions

Dynamic rheological behaviors of α-cellulose 1-ethyl-3-methylimidazolium acetate ([Emim]Ac) solutions were investigated in a large range of concentrations (0.1–10 wt %) at 25 °C. On the basis of data from the dynamic viscoelastic test, the exponents of the specific viscosity η _sp versus concentration c were determined as 1.0, 2.0 and 4.7 for dilute, semidilute unentangled and entangled regimes respectively, which were in accordance with the scaling prediction for neutral polymer in θ solvent. The intrinsic viscosity [η] of the solution was determined to be 253 mL/g at 25 °C. The linear viscoelastic response of the dilute and semidilute unentangled solutions could be described successfully by the Zimm and Rouse model (ν = 0.5 for θ solution) respectively, suggesting that the motion of cellulose chain in [Emim]Ac changed from Zimm to Rouse model with increasing concentration. At low concentrations, failure of the Cox–Merz rule with steady shear viscosity larger than complex viscosity was observed. While as the concentration increased, the deviation from the Cox–Merz rule disappeared due to the formation of homogeneous entanglement structure in cellulose solution.     

Effects of novel quasi-interpenetrating network/gold nanoparticles composite matrices on DNA sequencing performances by CE

Gold nanoparticles (GNPs) with particle sizes of about 20, 40, and 60 nm were prepared and added into a quasi-interpenetrating network (quasi-IPN) composed of linear polyacrylamide (LPA) with different viscosity-average molecular masses of 1.5, 3.3, and 6.5 MDa and poly-N,N-dimethylacrylamide (PDMA) to form polymer/metal composite matrices, respectively. These novel matrices could improve ssDNA sequencing performances due to interactions between GNPs and polymer chains and the formation of physical cross-linking points as demonstrated by intrinsic viscosities and glass transition temperatures. The effects of the parameters in relation to quasi-IPN/GNPs matrices, such as GNP contents, GNP particle sizes, LPA molecular masses, and solution concentrations, on ssDNA sequencing performances were studied. In the presence of GNPs, the separation had the advantages of high resolution, speediness, excellent reproducibility, long shelf life and easy automation. Therefore, less viscous matrix solutions (with moderate size GNPs) due to lower solution concentration and lower-molecular-mass LPA could be used to replace more viscous solutions (without GNPs) due to higher solution concentration or higher-molecular-mass LPA to separate DNA, while the sieving performances were approximate even higher, which helped to achieve full automation especial for capillary array electrophoresis (CAE) and microchip electrophoresis (MCE).     

Divergent dispersion behavior of ssDNA fragments during microchip electrophoresis in pDMA and LPA entangled polymer networks

Resolution of DNA fragments separated by electrophoresis in polymer solutions ("matrices") is determined by both the spacing between peaks and the width of the peaks. Prior research on the development of high-performance separation matrices has been focused primarily on optimizing DNA mobility and matrix selectivity, and gave less attention to peak broadening. Quantitative data are rare for peak broadening in systems in which high electric field strengths are used (>150 V/cm), which is surprising since capillary and microchip-based systems commonly run at these field strengths. Here, we report results for a study of band broadening behavior for ssDNA fragments on a glass microfluidic chip, for electric field strengths up to 320 V/cm. We compare dispersion coefficients obtained in a poly(N,N-dimethylacrylamide) (pDMA) separation matrix that was developed for chip-based DNA sequencing with a commercially available linear polyacrylamide (LPA) matrix commonly used in capillaries. Much larger DNA dispersion coefficients were measured in the LPA matrix as compared to the pDMA matrix, and the dependence of dispersion coefficient on DNA size and electric field strength were found to differ quite starkly in the two matrices. These observations lead us to propose that DNA migration mechanisms differ substantially in our custom pDMA matrix compared to the commercially available LPA matrix. We discuss the implications of these results in terms of developing optimal matrices for specific separation (microchip or capillary) platforms.     

Fluorescent-based single-strand conformation polymorphism/heteroduplex capillary electrophoretic mutation analysis of the P53 gene.

Fluorescent-based single-strand conformation polymorphism (F-SSCP) analysis with capillary electrophoresis (CE) is the most common method for the detection of mutation because of its high sensitivity and resolution. In this study, we prepared an inexpensive linear polyacrylamide (LPA), and successfully applied it to CE-SSCP analysis and tandem CE-SSCP/heteroduplex analysis (HA) of the P53 gene on an ABI capillary genetic analyzer. A comparison of the sieving capabilities of a homemade LPA and commercial polydimethylacrylamide (PDMA) demonstrates that the homemade LPA has a higher resolution, a shorter analysis time, and is more suitable for tandem SSCP/HA than commercial PDMA. To show the usefulness, mutations of P53 gene exon 7 - 8 in 37 tumor samples were investigated by using homemade LPA. The results indicate that 10 mutations were found in 9 of 37 cases; the majority of P53 mutations were missense mutations, and 70% were located in exon 7, which plays an important role in neoplastic progression in human tumorigenesis.     

Properties of star-branched and linear chains in confined space: a computer simulation study

We studied the properties of simple models of linear and star-branched polymer chains confined in a slit. The polymer chains were built of united atoms and were restricted to a simple cubic lattice. Two macromolecular architectures of the chain linear and star-branched with three branches (of equal length) were studied. The excluded volume was the only potential introduced into the model and thus, the system was athermal. The chains were put between two parallel and impenetrable surfaces. Monte Carlo simulations with a sampling algorithm based on chain’s local changes of conformation were carried out. The differences and similarities in the global size and the structure and of linear and star-branched chains were shown and discussed.     

Charge density effects in salt‐free polyelectrolyte solution rheology

Solution rheology of 2‐vinyl pyridine and N‐methyl‐2‐vinyl pyridinium chloride random copolymers in ethylene glycol was studied over wide ranges of concentration and effective charge. The fraction of quaternized monomers α and the fraction of monomers bearing an effective charge f of these copolymers were measured using counterion titration and dielectric spectroscopy, respectively. Ethylene glycol is a good solvent for neutral poly(2‐vinyl pyridine), with very few ionic impurities. The viscosity η and relaxation time τ of dilute and semidilute unentangled solutions exhibit the scaling with concentration and effective charge expected by the Dobrynin model. Reduced viscosity data are independent of concentration in dilute solution, giving an intrinsic viscosity that depends on effective charge, and the experimental data obey the Fuoss law in the semidilute unentangled regime. Scaling concentration with the overlap concentration (c/c*) reduces these data to common curves, and c* ～ f ^?12/7 as predicted by the Dobrynin model, where f is the fraction of monomers bearing an effective charge. While the overlap concentration depends strongly on effective charge until counterion condensation occurs, the entanglement concentration c_e is surprisingly insensitive to effective charge, indicating that entanglement effects are not understood using the Dobrynin model. The terminal modulus G = η/τ depends only on the number density of chains G = ckT/N for c* < c < c_e, and G ～ c^3/2 for c > c_e independent of the effective charge. © 2006 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 44: 2001–2013, 2006     

Capillary zone electrophoresis of proteins in semidilute polymer solutions: inter- and intra-polymer predictability of size-dependent retardation.

The retardation of three "spherical" proteins with Stokes' radii of 2.0, 2.4, and 3.0 nm (35-104 kDa) was studied in capillary zone electrophoresis (CZE), using semidilute solutions of polyethylene glycol (PEG), linear polyacrylamide (PA), and polyvinyl alcohol (PVA). The purpose was to test the models predicting that the ratio of particle radius, R, to the mesh size of polymer network (the correlation or screening length of a semidilute polymer solution), xi, directly governs the size-dependent retardation in the form: mu/muo = exp (-R/xi). Here xi = kc-0.75, where c is polymer concentration and the numerical factor kcan be calculated based on polymer molecular weight. In application to polymers in a "good solvent" (PA and PEG in the aqueous buffer) and to proteins of 2.4 and 3.0 nm radius, that relation between relative mobility and R/xi was found to be obeyed for PA, while for PEG the value of k derived from retardation experiments significantly exceeded that which was theoretically calculated. Thus, the retardation appears to be polymer-specific, rather than universal, even for polymers in a "good solvent". It is suggested that, in that case, retardation of proteins of R > 2 nm be quantitatively described in the form mu/muo = exp[-p(R/xi], where p is a parameter depending on monomer type and/or polymer polydispersity. For PVA, the logarithm of mu/muo was found to be linearly related to c (in line with the prediction that the aqueous buffer is a "poor solvent" for this polymer) and to be near-independent of R.     

A new quasi-interpenetrating network formed by poly(N-acryloyl-tris-(hydroxymethyl)aminomethane and polyvinylpyrrolidone: separation matrix for double-stranded DNA and single-stranded DNA fragments by capillary electrophoresis with UV detection

The preparation of a new separation matrix, quasi-interpenetrating networks (quasi-IPNs) formed by poly(N-acryloyl-Tris) (poly(tris-A)) and PVP, and its application for dsDNA and ssDNA fragments separation by CE with UV detection, are presented. This new quasi-IPN exhibited high sieving performance, good dynamic coating ability, and low viscosity. Single-base resolutions of dsDNA fragments (Rs = 0.92 for 123/124 bp) and ssDNA fragments (Rs = 0.65 for 123/124 base, Rs = 0.48 for 309/310 base) were achieved by using the quasi-IPN of poly(tris-A)/PVP (2% + 2%) solution in a 31 cm effective length linear polyacrylamide (LPA)-coated column. Single-base separation of dsDNA fragments (Rs = 0.92 for 123/124 bp) was also obtained within 28 min in a 46.7 cm effective length bare column at higher 160 V/cm electric field strength by using the same quasi-IPN solution. The RSD of the migration time measured for each DNA fragments was less than 1.5% in the bare column for nine continuous runs. The effects of temperature and electric field strength on the DNA separation were also investigated.