**2.1 The DNA-chitosan system**

Upon mixing, oppositely charged polyelectrolytes interact electrostatically and form complexes in a process that is promoted by an increase in entropy which is due to a release of counterions (Manning, 1978; Matulis et al., 2000). Accordingly, polycation molecular parameters such as charge density and valence have gained attention in recent reports (Danielsen et al., 2004; Maurstad et al., 2007).

The role of chitosan charge density is well established. It is accepted that the high charge density of chitosan at pHs below its pKa results beneficial for polyplex preparation, and also that its low charge density at pH 7.4 contributes to a low polyplex cytotoxicity and facilitates the intracellular release of DNA from the complex after its endocytotic cellular uptake (Strand et al., 2010). By contrast, the role of chitosan valence on transfection efficiency is contradictory. Namely, while several studies promote the use of high Mw chitosans (Huang et al., 2005; MacLaughlin et al., 1998), some other publications report that lower Mw chitosans are superior for gene transfer (Koping-Hoggard et al., 2003; Lavertu et al., 2006).

Aiming to draw general conclusions about the feasibility of using chitosan as a gene carrier, we characterized the DNA complexation and transfection mediated by three chitosans presenting different Mw (three different valences) at three different pHs of 5.0, 6.0, and 6.5 (three different charge densities). Table 1 summarizes the physical characteristics of the chitosans employed.


Table 1. Chitosans employed.
