Microalgae have been proposed for the bioremediation of wastewater, as well as for biofertilization and biostimulation of several plant species. This study used Limnospira sp. biomass produced in brewery wastewater to formulate a pelletized biofertilizer. Its efficacy in promoting barley (Hordeum vulgare) growth was then compared with chemical fertilizers and a control group without fertilization on lab- and pilot-scale setups. On a 100-day lab-scale experiment under controlled light (260–280 µmol photons m−2 s−1) and temperature (20 ± 2 °C), minor differences in plant growth were observed, whereas the elemental composition of the barley plants did not differ, including toxic elements. On a pilot-scale agricultural setup (5 m2) under environmental conditions, barley productivity, protein content, and the percentage of class I grains (diameter ≥ 2.5 mm) significantly increased based on the different dressing techniques used (p < 0.05). Using the microalgae-based biofertilizer for both base and top dressing increased productivity, protein content, and grain size (% class I) by 26.9%, 14.4%, and 8.78%, respectively, compared to using chemical fertilizers (NPK 5:20:20 and urea). These results indicate the great potential of using microalgae biomass from wastewater treatment as biofertilizer for more sustainable agriculture.