Hydrophilic nanocarriers, formed by electrostatic interaction between their oppositely charged components, are recognized as nanomaterials with an enormous potential as vectors in biomedical and pharmaceutical applications. However, comprehensive information about their fate in various biological environments is still lacking. In general, due to their size, nanoparticles are able to enter and transport molecules into cells via pathways forbidden for bigger structures of even identical chemical nature. Research under defined conditions aiming at a better understanding of nanoparticle-cell interactions is crucial for the development of safe and successful nanocarriers for biomedical and pharmaceutical applications. Nanogels produced by electrostatic interaction of two types of the polysaccharide chitosan and pentasodium triphosphate were the subject of the research of this thesis. Additional surface coating with sodium alginate conferred to the nanogels a negatively charged surface enabling sufficient stability under physiological and in vivo relevant conditions. Of particular interest was the comparison of the two chitosan types, the first one obtained from usual animal sources while the second one became newly available from fungal sources. Prerequisite for the comparison in terms of nanogel formation, stability, interaction with cells and potential applications was the detailed analysis and characterization of the chitosan samples. The identification of almost identical degree of deacetylation, very similar molar mass distribution and solution behavior, but a somewhat higher molar mass of the animal chitosan as the only difference, allowed for a direct comparison. The animal and fungal derived chitosan samples yielded similar nanogels in relation to formation, size distributions and zeta potential. The nanogels were spherical with a hydrodynamic diameter in the range of 450 nm and a zeta potential of approx. -65 mV. The physical characteristics of the nanogels were clearly influenced by the ionic strength and temperature of the suspension. High deformability of the nanogel network was demonstrated by filtration through pores much narrower than the gel diameter. Temperature cycles in the range of 4°C to 37°C proved reversibility of size and zeta potential. The nanogels were stable in both simulated in vivo environment and in conditioned cell culture media. Exposure of seven human cancerous and non-cancerous cell lines to the nanogels, to identify the influence on the cell metabolic activity, revealed cell-type, nanogel dosage and exposure time dependence. However, nanogels prepared from animal and fungal chitosan showed similar behavior in vitro in all cell lines tested. The results obtained so far confirm fungal derived chitosan as an alternative to the animal derived chitosan. Nanogels carrying model protein antigen cargoes were taken up by porcine monocyte-derived dendritic cells, and kinetic analysis using an auto-quenched protein revealed a time-dependent endocytic processing of the nanogel cargo. These in vitro analyses were elaborated with an in vivo murine immunization experiment. Nanogels carrying ovalbumin (OVA) induced a dose-dependent antibody response over 6 weeks post-vaccination. An effective memory response was induced in nanogel-OVA immunized mice following a 0.1 µg OVA boost injection. Nanogels were also prepared carrying recombinant vaccine antigen against Neospora caninum. These induced a protective immune response following both intranasal and intraperitoneal vaccination in mice challenged with the tachyzoite form of the parasite. Overall, the results obtained in the framework of this research extend the knowledge on chitosan-based nanogels intended as nanocarriers for biomedical/pharmaceutical applications and contribute to further development of effective nanogel-based delivery systems.