An Electrochemical Biosensor Sensitive to Structure Changes in an Absorbed Phospholipid Layer:

Development of a Phospholipase C Biosensor.

John T. Elliott
Biotechnology Division

We have fabricated an electrochemical biosensor for detecting structure changes in a phospholipid layer absorbed onto a mixed thioalkyl self-assembled monolayer (SAM). Mixed SAMs containing alkane terminated thiaethyleneoxide derivatives and b-mercaptoethanol were formed on gold surface electrodes. b-mercaptoethanol was used to provide pinhole defects that would allow access of a soluble redox species (K3Fe(CN)6) to the electrode surface. The addition of phospholipid vesicles (POPC) to the coated electrode surface resulted in a greater than 3-fold reduction in the observed faradaic current indicating the phospholipid layer limits access of the soluble redox species to the working electrode. Although in situ ellipsometry data suggests approximately one monolayer of phospholipid was deposited, impedance spectroscopy and the blocking layer formation kinetics indicate a more complicated phospholipid surface (e.g. a partial vesicle layer) may be present on some SAMs. To determine if structure changes in the phospholipid layer could be detected in cyclic voltammetry scans, we added phospholipase C (PLC) to the assembled electrode surface. Lipase activity resulted in an increase in the faradaic current to the level present before phospholipid coverage. This indicates that PLC caused structure defects in the blocking phospholipid layer that allowed the soluble redox species access to the working electrode. Further analysis of the phospholipid layer structure is required to establish factors that may improve sensitivity, reproducibility and stability of the sensor system.