Sol–gel nanoporous silica as substrate for immobilization of conjugated biomolecules for application as fluorescence resonance energy transfer (FRET) based biosensor

Abstract

Nanoporous silica beads with a particle size in the nanometer range, synthesized by adopting the sol–gel method, can be used as a dip-test platform for Förster Resonance Energy Transfer (FRET)-based sensing of troponin I, a regulatory protein for the contraction of cardiac muscles. The silica nanogel was synthesized using the sol–gel method, with tetraethoxysilane as the precursor, which was then subjected to hydrolysis in aqueous oxalic acid. The gel was molded and dried at 70°C, and subsequently calcined at 400°C. The silica substrate was then converted into a biosensor for troponin I by first silanizing, and then crosslinking Cy3 (FRET donor) labeled troponin I antibody (IgG) to which Cy5 (FRET acceptor) labeled Protein A was anchored. The potential of this biosensor to detect troponin I was examined using a combination of steady state fluorescence spectroscopy and microscopy. We found a considerable increase in the Cy3-to-Cy5 FRET efficiency as soon as troponin I was added to the biosensor. Results of this research show that the detection limit of troponin I using the FRET system is 10–5ngmL−1.