Electrospun natural biopolymers are of great interest in the field of regenerative medicine due to their unique structure, biocompatibility, and potential to support controlled release of bioactive agents and/or the growth of cells near a site of interest. The ability to electrospin chitosan and alginate to form polyionic complexed nanofibrous scaffolds was investigated. These nanofibers crosslink in situ during the electrospinning process, and thus do not require an additional chemical crosslinking step. Although poly(ethylene oxide) (PEO) is required for the electrospinning, it can be subsequently removed from the nanofibers simply by incubating in water for a few days, as confirmed by attenuated total reflectance Fourier transform infrared. Solutions that allowed uniform nanofiber formation were found to have viscosities in the range of 0.15-0.7 Pa·s and conductivities below 4 mS/cm for chitosan-PEO and below 2.2 mS/cm for alginate-PEO. The resultant nanofibers both before and after PEO extraction were found to be uniform and on the order of 100 nm as determined by scanning electron microscopy. The dynamic rheological properties of the polymer mixtures during gelation indicated that the hydrogel mixtures with low storage moduli provided uniform nanofiber formation without beaded structures. Increased amounts of chitosan in the PEO-extracted chitosan-alginate nanofibers resulted in a lower swelling ratio. Additionally, these nanofibrous scaffolds exhibit increased cell adhesion and proliferation compared to those made of alginate alone, due to the presence of the chitosan, which promotes the adsorption of serum proteins. Thus, these nanofibrous scaffolds formed purely via ionic complexation without toxic crosslinking agents have great potential for guiding cell behavior in tissue regeneration applications.