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Fig. 4 Number of particles per l L versus cation/anion ratio, for combinations of different polycations with a 250 kDa and b 700 kDa CMC
Overall, the changes in the determined SSC values were quite small as function of cation/anion ratio for all of these PEC mixtures. Larger and more noticeable changes took place in the average light scattered in forward direction (FSC) of the PEC mixtures. The FSC of PECs formed from polyDADMAC and CMC decreased dramatically as the cation/anion ratio increased between 0.5 and 2, even though this type of behavior was not reflected in the determined SSC values (Fig. 5b). The FSC for PECs formed from Raifix 25035 and CMC also decreased noticeably after the theoretical neutraliza- tion point. The FSC of PECs from Raisabond 15 and Raifix 01015 SW were overall significantly higher than for the other mixtures. Large differences in the FSC values as a function of cation/anion ratio, that were not reflected in the SSC values, indicated that FSC and SSC were connected to very different properties of the PEC particles. Available explana- tions from medical science about FCM of cells may be useful for understanding the scattered light in different directions of these PEC particles. In medical science, SSC is commonly used as an indicator of the granularity of cells, i.e. the amount of granular components within a cell that are able to scatter light in the side direction (Shapiro 2003). Therefore, SSC is not a direct measurement of particle size, but can be used to distinguish cell types from each other. Degranulation of cells and membrane ruffling following activation are known to decrease the SSC of cells. On the other hand, FSC is commonly used as an indicator of cell size, but is not a direct measurement of cell size (Shapiro 2003). It is known that many factors influence the FSC of a cell, such as
increasing the molar mass of cationic starch results in a larger hydrodynamic thickness of the adsorbed layer on quartz surfaces (Tammelin et al. 2004). The M w of PEC particles have previously been shown to increase with increasing molar mass of the used polyDAD- MAC (Dautzenberg 2000). It would be expected that increasing the molar mass of CMC in these experi- ments also would lead to thicker, more compact particles in suspension. It is important to remember that particles smaller than 0.1 l m were not analyzed in the FCM measure- ments, and are not included in the results. This phenomenon is highlighted when combining Raifix 01015 SW with 250 kDa CMC at a cation/anion ratio of 5:1, which resulted in a coagulating combination of polyelectrolytes (Fig. 4a). Close to the neutralization point, the large number of particles below the detec- tion limit were suddenly elevated into the detection range of the FCM, which resulted in a large spike in particles per l L.
Light scattering properties
PEC mixtures were analyzed by flow cytometry at different cation/anion ratios. The scattering of light in side direction (SSC), and scattering of light in forward direction (FSC) was recorded for the particles. Aver- age values were calculated for the different PEC populations, and these values were plotted as a function of cation/anion ratio. Mixing together poly- DADMAC with CMC resulted in PEC particles with quite high SSC values (Fig. 5a). The SSC of particles consisting of cationic starches and CMC were con- siderably lower at all measured cation/anion ratios.
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