A new approach for electric field gradient focusing (EFGF) based on the use of a variable-width polyaniline (PANI) electrode is demonstrated. The electrode was created by patterning a PANI nanofibre film using a 635-nm laser and a computer-controlled XY stage. The electrode consisted of 16 segments of varying width, ranging from 200 to 5000 mu m in 320 mu m increments, with the resistance changing approximately 20-fold from 3881 to 198 k Omega at each extreme, respectively. Application of a voltage across the electrode established a voltage gradient resulting in a non-linear distribution of electrophoretic velocities along the microchannel. When balanced with a combination of hydrodynamic flow and EOF, the variable-width PANI electrode could be used for the concentration and separation of two cationic dyes, rhodamine 6G and quinine, which were concentrated by at least threefold within 10 min.