Abstract The influence of sodium alginate sterilization on the viability and mitotic activity of embedded protoplasts was studied in protoplasts of Brassica oleracea subsp. alba and rubra isolated from hypocotyl tissue and leaves of seedlings or plants grown in vitro. Both leaf and hypocotyl-derived protoplasts were more viable and divided more frequently when embedded in filtrated alginate. Division frequency was highest in cv. Reball F1 and the mitotic activity of its protoplasts was three times higher when embedded in filtrated alginate (36.1 ± 6.8%) than when cultured in autoclaved alginate (10.9 ± 5.0%). Protoplast-derived calli colonies were transferred to solid regeneration media and plants of all tested accessions were obtained.