Novel renewable immunosensors based on temperature-sensitive PNIPAAm bioconjugates

A novel renewable immunosensor was created comprising a temperature-controlled surface composed of poly(n-isopropylacrylamide) (PNIPAAm)–antibody conjugates that could reversibly bind the antigen. Bovine serum albumin (BSA) and the corresponding antibody (anti-BSA) were chosen as a model antibody–antigen system to demonstrate the concept. The thermally responsive PNIPAAm conjugated to anti-BSA displayed a controllable conformation change between an expanded and a collapsed form, below and above its characteristic phase transition temperature, i.e. low critical solution temperature (LCST). This showed a remarkable change in the bioaffinity of the conjugate for BSA. Thus, a renewable anti-BSA surface was generated for re-binding of the target antigen at the thermally controllable PNIPAAm–anti-BSA conjugated surface. The temperature-controlling strategy resulted in the regeneration of immunosensors on which immobilized anti-BSA antibodies retained their activity and specificity for more than 30 reproducible assays. The level of dissociation reached 89%, which is comparable with established recovery methods, while offering easer handing. The controlled binding and dissociation were monitored by quartz crystal microbalance (QCM), confocal fluorescence, native electrophoresis, laser-induced fluorescence, and electrochemical impedance methods.