Viable tomato mesophyll protoplasts were isolated by using the non-phytotoxic macerating enzyme PATE instead of the generally used Macerozyme. To obtain the osmotic conditions required during isolation, salt solutions, containing 2.5% (w/v) KC1 and 1% (w/v) MgS04 ■ 7H20, were used. From studies in different culture media it appeared that, under optimum conditions, cells regenerated a wall and divided three to four times. After about three weeks of incubation cell division stopped and cells started to degenerate.