Generation of natural pH gradients in microfluidic channels for use in isoelectric focusing

As a part of an ongoing investigation of the use of isoelectric focusing (IEF) in microfluidic devices, pH gradients were electrochemically formed and optically quantified in microfluidic channels using acid-base indicators. The microchannels consisted of two parallel 40-mm-long electrodes with an interelectrode gap of 2.54 mm; top and bottom transparent windows were separated by 0.2 mm. Gradients in pH were formed as a result of the electrochemical decomposition of water at an applied potential not higher than 2.5 V to avoid generation of gas bubbles. Solutions contained low concentrations of a single buffer. The stability of the pH gradients and their sensitivity to changes in initial conditions were investigated under static (nonflow) conditions. Isoelectric focusing of sample biological analytes, bovine hemoglobin and bovine serum albumin, was performed to illustrate the potential of "microfluidic transverse IEF" for use in continuous concentration and separation systems.