Nanoparticles were washed three times with milli-Q, and the nanoparticles collected after each washing step by centrifugation using a Heraeus Megafuge 1.0 R centrifuge (Thermo Scientific) for 30 min at 23,000 × g. The particles were redispersed in water prior to lyophilization. Nanoparticle formulations intended to have external lyoprotectant were redispersed in water containing trehalose, glucose, sucrose, fructose and sorbitol at a concentration of 10% (w/v). The nanoparticles were poured into semi-stoppered glass vials with slotted rubber closures at a maximal height of 10 mm. All formulations were lyophilized in a VirTis Advantage Plus Benchtop lyophilizer (SP Scientific, Warminster, PA, USA), by freezing at −55 °C for 5 h, followed by a first drying step at −45 °C and pressure of 400 mTorr during 40 h, followed by a secondary drying step at 20 °C and pressure of 200 mTorr during 8 h.