This study investigated the extraction and colloidal properties of pea protein isolates (PPIs) from four pea cultivars produced in a southern region in Italy. The control PPIs (C-PPIs) were obtained via an alkaline extraction and isoelectric precipitation method and then subjected to ultrasound treatment (US) to yield US-PPIs. The effects of cultivars and sonication on the physicochemical characteristics and emulsifying property of the PPIs were studied. Fourier-transform infrared (FTIR) spectroscopy, colorimetric measurement, dynamic light scattering (DLS), and confocal laser scanning microscopy (CLSM) were applied to characterize the samples. DLS results showed that C-PPIs displayed smaller particle sizes (8.86-15.9 µm) in comparison to US-PPIs (15.8-66.5 µm). DSC data showed that US-PPIs had improved thermal stability compared to control PPIs. FTIR analysis detected differences in the protein secondary structure of the various cultivars and between the native and US-PPIs. Emulsion stability studies indicated that emulsions stabilized with C-PPIs exhibited lower droplet sizes, implying improved stability in comparison to emulsions stabilized with US-PPIs. In conclusion, PPIs can be successfully extracted from different cultivars and applied as a natural emulsifier.