A mixture of nanospheres and nanoplates was synthesized through bioreduction of HAuCl4 with Cacumen Plarycladi leaf extract. In this work, shape separation of the nanospheres and nanoplates was achieved via agarose gel electrophoresis (AGE). Furthermore, the separation mechanism and the effects of parameters, including the agarose gel concentration, electrophoresis voltage and TBE buffer concentration, on the separation performance were investigated. The optimum conditions were found to be 1.2% gel concentration, 100 V voltage, and 1 x TBE buffer solution. Gold nanoparticles (NPs) prepared via a biological method were compared with those synthesized using a chemical method in the electrophoresis process with different characteristics. The results showed that the particles prepared via the biological method had good stability in a buffered solution without further modification. (C) 2015 Elsevier B.V. All rights reserved.