Fabrication of immuno-assay based diagnostic devices via controlled protein immobilization on amino-terminated organic films on silicon substrates for applications in forensics

The controlled immobilization of proteins on solid substrates has been extensively studied due to its application in many areas of diagnostics including forensics. Amino-terminated organic films were prepared on silicon wafers by self-assembling 3-aminopropyltriethoxysilane (APTES). Surface amino groups were derivatized into three different linkers; N-hydroxysuccinimide (NHS) ester, hydrazide, or maleimide ester groups. Protein immobilizations were carried out on modified APTES films containing these linkers via coupling with primary amines (–NH2) in intact monoclonal rabbit immunoglobulin G (IgG), the aldehyde (–CHO) of an oxidized carbohydrate residue in IgG, or the sulfhydryl (–SH) of fragmented half-IgG, respectively. Biological activity and long-term hydrolytic stability of immobilized IgGs were estimated by fluorescence measurements of adsorbed antigen. Our results indicate that the FITC-Ab binding capacity of the half-IgG immobilized via maleimide groups is greater than that of the oxidized IgG and the intact IgG immobilized via hydrazide and NHS ester groups.