An innovative approach to imprinted nanoparticles (nanoMIPs) is represented by solid-phase synthesis. Since the polymeric chains grow over time and rearrange themselves around the template, the binding properties of nanoMIPs could depend on the polymerization time. Here we present an explorative study about the effect of different polymerization times on the binding properties of ciprofloxacin-imprinted nanoMIPs. The binding properties towards ciprofloxacin were studied by measuring the binding affinity constants (Keq) and the kinetic rate constants (kd, ka). Furthermore, selectivity and nonspecific binding were valued by measuring the rebinding of levofloxacin onto ciprofloxacin-imprinted nanoMIPs and ciprofloxacin onto diclofenac-imprinted nanoMIPs, respectively. The results show that different polymerization times produce nanoMIPs with different binding properties: short polymerization times (15 min) produced nanoMIPs with high binding affinity but low selectivity (Keq > 107 mol L−1, α ≈ 1); medium polymerization times (30 min–2 h) produced nanoMIPs with high binding affinity and selectivity (Keq ≥ 106 mol L−1, α < 1); and long polymerization times (>2 h) produced nanoMIPs with low binding affinity, fast dissociation kinetics and low selectivity (Keq ≤ 106 mol L−1, kdis > 0.2 min−1, α ≈ 1). The results can be explained as the combined effect of rearrangement and progressive stiffening of the polymer chains around the template molecules.