We report here a new methodology for sorting mammalian cells based on their intrinsic motility on planar substrates, independent of chemoattractants and external fields. This biological analogue of thin layer chromatography consists of arrays of asymmetric adhesive islands on tissue culture dishes that rectify the random movement of cells and direct their migration in a specific direction. We demonstrated the use of planar cell chromatography in the separation of mixtures of 3T3 fibroblasts that express constitutively active Rac1 or RhoA and mixtures of 3T3 fibroblasts and SH-SY5Y neuroblastoma cells.