Light scattering concentration dependence

Ferri, F., Greco, M., Arcovito, G., De Spirito, M., and Rocco, M. (2002). Structure of fibrin gels studied by elastic light scattering techniques Dependence of fractal dimension, gel crossover length, fiber diameter, and fiber density on monomer concentration. Phys. Rev. E. 66(1 Pt 1), 011913. 

FP assays are known to be susceptible to artifacts (Turek-Etienne and Small, 2003). In principle, the assays are ratiometric and should normalize for variations in total excitation energy applied as would occur with inner filter effects, and newer generations of red-shifted fluorophores should help to eliminate interference (Vedvik et al., 2004). However, introducing a test compound with fluorescent or absorbent properties at 5 or 10 pM with the typically sub-micromolar concentrations of fluorophores in an FP assay can significantly skew the measurements. For example, if the compounds are insoluble, they can scatter and depolarize light. A concentration-dependent effect on an FP assay could result from an increase in the amount of insoluble compound. 

Thus we have finally established how light scattering can be used to measure the molecular weight of a solute. The concentration dependence of r enters Eq. (10.54) through an expression for osmotic pressure, and this surprising connection deserves some additional comments ... 

The increase in Ca is initiated rapidly and begins to recover after 1 min. The order of potency correlates fairly well with the solubilities of these compounds in organic solvents (37) and their abilities to accumulate in phospholipid vesicles (38), i.e., 6>y>a>p, but not with their insecticidal activity (y 6>a p 39). At these concentrations, crystals of p-, a-, and y-HCH were evident in the cell suspensions when we made simultaneous measurements of the right-angle light scatter, indicating that the order of aqueous solubilities is 6>y>a>p. However, stimulation by 6-HCH at concentrations below its aqueous solubility limit shows a typical dose dependency of the response (Figure 10). 

The various physical methods in use at present involve measurements, respectively, of osmotic pressure, light scattering, sedimentation equilibrium, sedimentation velocity in conjunction with diffusion, or solution viscosity. All except the last mentioned are absolute methods. Each requires extrapolation to infinite dilution for rigorous fulfillment of the requirements of theory. These various physical methods depend basically on evaluation of the thermodynamic properties of the solution (i.e., the change in free energy due to the presence of polymer molecules) or of the kinetic behavior (i.e., frictional coefficient or viscosity increment), or of a combination of the two. Polymer solutions usually exhibit deviations from their limiting infinite dilution behavior at remarkably low concentrations. Hence one is obliged not only to conduct the experiments at low concentrations but also to extrapolate to infinite dilution from measurements made at the lowest experimentally feasible concentrations. 

Thus both the numerator and denominator terms in Eq. (41), or in Eq. (44), depend on the concentration. Because of this situation empirical extrapolation of D is particularly hazardous (for random coiling polymers). If F2 is known from osmotic or light-scattering measurements at a series of concentrations, extrapolation according to Eq. (44) will be facilitated. (If such measurements have been carried out, however, the molecular weight also will have been determined.)... 

This expression can be modified to apply directly to any of various techniques used to measure the interaction parameter, including membrane and vapor osmometry, freezing point depression, light scattering, viscometry, and inverse gas chromatography [89], A polynomial curve fit is typically used for the concentration dependence of %, while the temperature dependence can usually be fit over a limited temperature range to the form [47]... 

The lifetime detection techniques are self-referenced in a sense that fluorescence decay is one of the characteristics of the emitter and of its environment and does not depend upon its concentration. Moreover, the results are not sensitive to optical parameters of the instrument, so that the attenuation of the signal in the optical path does not distort it. The light scattering produces also much lesser problems, since the scattered light decays on a very fast time scale and does not interfere with fluorescence decay observed at longer times. 

Equation 3.27 forms the basis for determination of Molecular weight from light scattering data. Like Osmotic pressure measurements, it is essential to consider the non-ideality of solutions and the concentration dependence. Following Debye, eq. 3.27 gets modified to... 

Properties which depend on the size of the chain, for example, viscosity and angular dependence of light scattering, get strongly affected by chain expansion. The viscosity may even increase markedly as polymer concentration decreases, the increase in viscosity results in an increase in the degree of... 

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