ABSTRACT
Improving the way by which proteins are immobilized on surfaces such that they maintain their functionality is of critical importance in medical diagnostics, bioreactors, and biosensors. Depending on the intended applications, proteins can be immobilized on surfaces by different approaches [1,2]. Immobilization through physical adsorption, though simple, usually has the problem of leakage of the immobilized proteins from the support and leads to partial denaturation of the protein. This results in a short lifetime of devices based on immobilized proteins and may even cause sample contamination, which is a great concern if these devices are to be used in vivo. Chemical immobilization methods eliminate the leakage problem by covalently tethering protein molecules to supports. Different types of functional groups, such as amino, sulfhydryl, and carboxyl groups, are frequently used in chemical immobilization of proteins to the support. Since each protein molecule usually has many of these functional groups distributed on its surface, attachment through these groups inevitably leads to a different orientation of the immobilized proteins on the support (hereafter referred to as random immobilization). In random immobilization, the active/binding site of the protein molecule may be partially or totally blocked by neighboring protein molecules or the immobilization surface due to mixed orientation of the immobilized protein (Fig. 1A). This is one of the reasons that chemical
immobilization often leads to a significant decrease in activity/binding capacity of the immobilized protein [3,4].
