The applicability of free flow electrophoresis (FFE) was expanded towards processing of sensitive cells. The chamber medium was adjusted to a physiologic pH of 7.35 by a mixture of N-(2-hydroxyethyl)piperazine-N'-(3-propanesulfonic acid) (EPPS) and 2,2-bis(hydroxymethyl)-2,2'2"-nitrilotriethanol (BISTRIS). These substances proved to be nontoxic to sensitive cells such as human smooth muscle or thyroid cells. They enhanced the electrical conductivity of the medium only slightly so that a new cell electrophoresis separation medium could be prepared, which contained 30 mM NaCl together with or without 1 mM CaCl2 but did not generate problems of overheating the fluid. Suspended in this medium, human smooth muscle cells as well as human thyroid carcinoma cells remained viable single cells for at least 120 min. After this period they could be recultured to form monolayers. if electrophoresed in the Octopus preparative FFE device, they migrated as single cells and did not clot; therefore, their electrophoretic behavior could be determined exactly.