Spherical and rod-shaped Au nanoparticles with various particle sizes (< 70 nm) were synthesized via the seeded growth method and utilized in gel electrophoresis under varied gel mesh sizes to investigate their migration velocities. Due to the strong effect of the gel, which consists of a three-dimensional mesh network with adjustable mesh sizes via the agarose concentration, the migration of the nanoparticles was retarded by steric and/or hydrodynamic interactions leading to e.g. friction as well as attraction and finally to decreased migration velocities. It was observed that the migration velocity of rod-shaped nanoparticles was less diminished in gels with smaller mesh sizes compared to spherical nanoparticles with one similar dimension (length of rod corresponded to diameter of sphere). These experimental data were used to validate modeled migration velocities using Henry’s equation. However, this retardation is not sufficiently addressed by Henry’s equation and therefore, a migration factor was included in the model, setting the mesh size in ratio to the particle size as well as the particle morphology. Despite its simplicity, this new model allows a more accurate prediction of the migration velocity during the gel electrophoresis under various conditions.