Lipid bilayer on a microdroplet integrated with a patterned Ag/AgCl microelectrode for voltage-clamp fluorometry of membrane transport

Abstract Voltage-clamp fluorometry (VCF) has been combined with artificial lipid bilayer systems to optically measure the dynamics and contribution of membrane proteins on molecular transport at the clamped membrane potential. However, the previous methods for VCF require expensive apparatus, such as total internal reflection fluorescence microscope, or require extensive experience for voltage-clamping, or possibly lead to imprecise membrane potential. This study describes VCF performed using a pico-liter-sized lipid bilayer chamber with an embedded Ag/AgCl microelectrode. Owing to the size of the picoliter order, molecular transport through the lipid bilayer can be detected using a normal fluorescent microscope. Easy electrical access into the lipid bilayer chamber can be realized because the microelectrode is spontaneously embedded inside when forming the lipid bilayer chamber. The microelectrode is made of Ag/AgCl; therefore, it does not tend to polarize, which results in reliable regulation of the membrane potential. Using the developed system, we performed the VCF of molecular transport through the lipid bilayer by incorporating the pore-forming membrane protein, α-hemolysin. This proposed method will increase the number of researches that can perform the VCF of molecular transport through artificial lipid bilayers.