Polyelectrolyte multilayers (PEMs) have been prepared using the strong polycation poly(diallyldimethylammonium chloride) and the weak polyanion poly(ethylene-alt-maleic acid) [P(E-alt-MA)]. The spin label (SL) 4-amino-2,2,6,6-tetramethylpiperidine 1-oxide was covalently attached to the polyanion [SL-P(E-alt-MA)] to monitor changes in the signal intensity as a measure of the layer growth and the line shape of the spectra indicative of the polymer chain mobility obtained by continuous wave electron paramagnetic resonance spectroscopy. The aim of this study was to investigate the influence of the preparation conditions. We show the influence of the pH value on the buildup and the mobility of PEMs with 16 and 17 layers, respectively. In the first part, the spin-labeled polyanion was used in the preparation of each polyanion layer to monitor the growth of the multilayers. For all pH values from 3 to 6 a non-linear growth behavior was obtained at the beginning of the PEM buildup where the surface is not yet fully covered, leading to a linear growth after reaching a transition point. This point depends on the pH value. In a second study, the spin-labeled polyanion was selectively placed in one layer to display changes in the rotational mobility of the SL in this layer within the multilayers. As a result, an increase of the rotational mobility of SL-P(E-alt-MA) with increasing pH value was observed.