Brassinosteroids, such as 24-epibrassinolide (EBR), are some of the biologically most active growth regulators that specifically modulate plant responses to abiotic stress. In this study, the ameliorative effect of EBR on growth, protein contents, and antioxidant enzymes was investigated in two Solanum tuberosum L. cultivars, Cardinal and Desiree, growing in vitro under salinity stress. EBR (0, 1, or 2 μM) was exogenously applied in two ways, i.e., EBR pretreatment of nodal explants for 8 h (PT) and addition of EBR directly into the Murashige and Skoog (MS) basal medium (IM) followed by explant inoculation. These explants were subjected to salt stress (0, 40, 60, or 80 mM) for 30 d. The growth of plants subjected to NaCl stress was substantially reduced. EBR pretreatment (both PT and IM) alleviated the harmful effects of salt stress for all the measured morphological and biochemical parameters. In general, the quantity of total soluble proteins, superoxide dismutase (SOD), and peroxidase (POD) increased in plants treated with EBR or NaCl alone but were reduced when both were supplemented together. Therefore, exogenous application of EBR not only played a role in terms of in vitro potato growth but also significantly affected the tested biochemical parameters.