To address targeting and bioavailability issues of peptidic drugs like desmopressin, the encapsulation into nanoparticles (NP) has become standard in pharmaceutics. This study investigated the encapsulation of desmopressin into PLGA NP by the use of pharmaceutically common stabilizers as a precursor to future, optional targeting and bioavailability experiments. Polymer dry weights were measured by freeze drying and thermo gravimetric analysis (TGA). Particle sizes (ranging between 105 and 130 nm, PDI < 0.1) and zeta potentials (−35 to −45 mV) were analyzed with Dynamic Light Scattering (DLS) and Laser-Doppler-Anemometry (LDA) respectively. Highest loading efficiencies, quantified by RP-HPLC, were achieved with Pluronic F-68 as stabilizer of the inner aqueous phase (1.16 ± 0.07 μg desmopressin/mg PLGA) and were significantly higher than coating approaches and approaches without stabilizer (0.74 ± 0.01 μg/mg). Optimized nanoformulations are thus in competition with the concentration of commercial non-nanoparticulate desmopressin products. Stability of desmopressin after the process was evaluated by HPLC peak purity analysis (diode array detector) and by mass spectrometry. Desmopressin was shown to remain intact during the whole process; however, despite these very good results the encapsulation efficiency turned out to be a bottle neck and makes the system a challenge for potential applications.