text: The DNA or RNA aptamers are fixed on the interrogating electrode, where a redox reaction is reported by a redox tag. Gold is often used as the probe surface for interrogating electrodes. The surface of the gold electrode is packed with redox-tagged DNA or RNA aptamers. The redox reporter is often methylene blue. Upon target binding, the aptamer changes structure by folding, bringing the redox reporter closer to the gold electrode. This increase in proximity from the redox-reporter to the electrode enables faster electron transfer from the redox tag to the gold electrode. The increase in speed of electron transfer contributes to a change in Faradaic current that is detected by the potentiostat.
  The reference electrode is the site of a known chemical reaction that has a known redox potential. For example, a reference electrode that harbors the reaction of silver-silver chloride (Ag/AgCl) has a fixed redox potential and is the measuring point for the redox potential of the interrogating electrode. The counter electrode (or auxiliary electrode) acts as a cathode or anode to the interrogating electrode. The applied voltage is not passed through the reference electrode due to an impedance supplied by the potentiostat. Therefore, the potential generated within the electrochemical cell is attributed to the interrogating electrode. Current is measured as potential of the interrogating electrode vs. the fixed potential of the reference electrode. The difference in potential is what produces the current in the external circuit and generates a signal. The signal quantifies target binding depending on electron transfer that is stoichimetrically proportional to target binding.
  Four electrode method has also been demonstrated in an electrochemical nanoporous alumina membrane sensor, where the aptamer was grafted onto the membrane and not on the electrode. The binding of the aptamer with the target protein produces a change in impedance of the membrane which is picked up by the electrochemical sensor using an impedance spectroscopy analyzer. This approach could be beneficial in cases where the electric field of the electrode can change the aptamer structure or the biointerface which may decrease the sensing ability.