Mixing oppositely charged polyelectrolytes and multivalent counterion solutions at low concentrations leads to the formation of colloidal ionically crosslinked polyelectrolyte particles. Due to the rapid reaction kinetics, the complexation processes and the final product could vary significantly when changing the mixing efficiency, which was often overlooked in previous studies. To investigate the effect of mixing on the polyelectrolyte-based colloid formation, we use chitosan/tripolyphosphate mixtures as a model system and compare the particle formation under flash nano-complexation (FNC, representing rapid and efficient mixing) and conventional dropwise mixing. It turns out that the non-uniform mixing and rapid complex formation during conventional mixing lead to particle formation at a low tripolyphosphate : chitosan ratio, which could be avoided by FNC. When mixing using FNC, the particle formation started at a critical tripolyphosphate : glucosamine ratio, below which only soluble complexes exist, and such a critical ratio is independent of the chitosan molecular weight and charge density. Homogeneous mixing also leads to the formation of a large amount of small primary particles without further aggregation due to the rapid consumption of free crosslinking counterions. Such a strong dependency of ionically crosslinked polyelectrolyte colloid formation on the mixing efficiency was also demonstrated using other polyelectrolytes and counterions. Thus, the mixing efficiency could have a significant impact on the interpretation of the complexation process and the mechanisms and should be carefully discussed when studying ionically crosslinked polyelectrolyte colloids.