동적광산란 기준 시료 측정 방법-2 : TEM과 DLS 입도 비교하기

Measuring Latex Standards

                                by Dynamic Light Scattering

Comparison of sizes measured by TEM and DLS

Different measurement techniques will measure different properties of a particle and so can give different results in terms of the size interpreted from the measured property.

Therefore, the question which often arises, is which one is the correct result? For many people ‘seeing is believing’ so the electron microscope result is ‘correct’. In fact, samples prepared for electron microscope examination are often harshly treated and this treatment can distort soft materials such as polymer lattices and change, or mask, surface structures.

It can make the size measurement of some types of materials like surfactant micelles impossible. In contrast, DLS measures the hydrodynamic diameter of dispersed particles in their native environment.

Any surface structure such as an adsorbed polymer layer, or a change in the electrical double layer that affects the Brownian motion of the particle, will change the effective particle size [5].

Increasing the surface structure or extending the electrical double layer by using a very low salt dispersant, will reduce Brownian motion and increase the measured size. For these reasons, the hydrodynamic size or DLS size of particles that are not smooth, hard  spheres is usually larger than the TEM size.

Sample Preparation of Latex Standards for DLS Measurement

For instruments with a 90o detection angle (for example, Zetasizer Nano S90, μV or APS), all latex standards are supplied at a concentration that is too high for DLS measurement, typically 1%w/v. Too concentrated a sample will produce multiple scattering resulting in the apparent size being too small. Therefore, the latex dispersion should be diluted with 10mM NaCl as previously discussed which has been filtered at 0.2 microns.

For backscatter detection instruments which have variable measurement positions (for example, Zetasizer Nano S or ZS), measurements are possible on neat concentrations of latex. The reduction in path length arising from the variable measurement position results in a reduction in multiple scattering.

However, the final concentration of any latex used should be such that the result is independent of the actual concentration [2,3] with the optimum concentration being dependent upon size of the latex.

Various Thermo Scientific Nanosphere 3000, 2000 and 4000 series size standards were prepared accordingly and measured on a range of Zetasizer instruments for this application note. Some of these standards were measured at neat concentration, others were diluted in 10mM NaCl and others were prepared in 13% w/v sucrose solution.

The density of polystyrene latex spheres is close to that of a 13% w/v sucrose solution and reduces problems of sedimentation of the latex. The concentration and diluent used for the various latex standards used in this application note are shown in Table 1.

Results and Discussion Table 1 summarises the results obtained for various latex standards measured by DLS. The table includes details of the latex used (with the Thermo part number displayed in brackets), the certified (#) or hydrodynamic (*) size range, the concentration at which the latex was measured, the diluent used for preparation, the instrument on which the measurements were taken and the z-average diameters obtained for each latex.

All of the standards available from Thermo have a certified size range but some also have hydrodynamic size ranges quoted. The certified size range (#) is obtained using transmission electron microscopy and is traceable to NIST. The hydrodynamic size range (*) is determined by DLS.

The results show that there is a wide concentration range over which latex standards can be measured. The backscatter detection used in the Zetasizer Nano allows some latex samples to be measured at neat concentration i.e. 1% w/v. As the size of the latex particles increase, the issue of number fluctuations and sedimentation become important.

During a DLS measurement, the intensity of scattered light fluctuates due to the Brownian motion of the particles. The scattering intensity is proportional to the sample concentration and so the number of particles within the scattering volume should remain constant during the course of the measurement. However, as the particle size increases, the number of particles in the scattering volume decreases until severe fluctuations of the momentary number of particles in the scattering volume will occur.

Number fluctuations are defined as variations in the number of particles within the scattering volume during the course of a DLS measurement. In order to avoid number fluctuations, the concentration of the sample must be increased.

However, this will increase multiple scattering effects which in turn will influence the result obtained. Back scatter detection, in combination with a variable measurement position, allows for higher sample concentrations to be measured thereby avoiding number fluctuation problems. Number fluctuations normally manifest themselves by elevated and fluctuating baselines in the correlation functions.

A second problem when measuring large sized particles by DLS is that of sedimentation. All particles will sediment and the rate will depend upon the particle size and relative densities of the particles and suspending medium. For DLS, the rate of sedimentation should be much slower than the rate of diffusion. Large particles diffuse slowly so sedimentation is a more important issue.

The presence of sedimentation can be determined by checking the stability of the count rate from repeat measurements of the same sample.

Count rates which are decreasing with successive measurements indicate that sedimentation is present and the Expert Advice system in the Zetasizer software will highlight this to the user.

It may be advantageous to suspend the particles in a medium of similar density if the viscosity is not significantly increased. For the measurements of 3, 6 and 8.9μm latex in this application note, the samples were prepared in 13% w/v sucrose which has the same density as the latex.

The results obtained for the 3, 6 and 8.9μm latex samples are in the expected size range using the DLS technique. They were measured at concentrations between 0.15 and 0.24% w/v. These results confirm that multiple scattering effects were minimised using back scatter detection and that number fluctuations and sedimentation did not influence the results obtained.

References
[1] R. Pecora (1985) Dynamic Light Scattering: Applications of Photon Correlation Spectroscopy. Plenum Press, New York.
[2] International Standard ISO13321 (1996) Methods for Determination of Particle Size Distribution Part 8: Photon Correlation Spectroscopy. International Organization for Standardization (ISO).
[3] International Standard ISO22412 (2008) Particle Size Analysis: Dynamic Light Scattering (DLS). International Organization for Standardization (ISO).

[4] National Institute of Standards and Technology (www.nist.gov).
[5] R.S. Chow and K. Takamura (1988) J. Colloid. Int. Sci, 125, 266.