Study of the Enzyme Activity Change due to Inkjet Printing for Biosensor Fabrication

Enzymes, the most commonly used biosensing element, have a great influence on the performance of biosensors. Recently, drop-on-demand (DOD) printing technique has been widely employed for the fabrication of biosensors due to its merits of noncontact, less waste, and rapid deposition. However, enzyme printing studies were rarely conducted on the effect of printing parameters from the aspect of the pressure wave propagation mechanism. This study investigated the effects of pressure wave propagation on enzyme activity from the aspects of wave superposition, wave amplitude, resulting mechanical stress, and protein conformation change using pyruvate oxidase as the model enzyme. We found that the mechanical stress increased the activity of pyruvate oxidase during the inkjet printing process. A shear rate of 3 × 10⁵ s^-1 enhanced the activity by 14.10%. The enhancement mechanism was investigated, and the mechanical activation or mild proteolysis was found to change the conformation of pyruvate oxidase and improve its activity. This study is fundamental to understand the effect of both printing mechanism and induced mechanical stress on the properties of biomolecules and plays an important role in modulating the activity of other enzyme-based inks, which is crucial for the development of biosensors.