One-step and high-density protein immobilization on epoxysilane-modified silica nanoparticles

Silica nanoparticles are most commonly modified with amino-silanes, followed by post-modification activation for protein immobilization. In this work, epoxy-functionalized silica nanoparticles were prepared by modification with glycidyloxypropyl trimethoxysilane (GPTMS) for direct protein immobilization. Silica nanoparticles possessed an average size of 46 nm, but increased to 63 nm after GPTMS modification. Reaction time, reaction temperature and GPTMS content had no significant effect on particle size. Zeta potential of SiO₂ changed from −26mV to +38mV after modification. Fourier-transformed infrared spectroscopy revealed alkyl C-H bending and stretching bands at 2944 cm⁻¹, 1343 cm⁻¹ and 1465 cm⁻¹, respectively, for the modified nanoparticles. Fluorescein cadaverine was found to bind to GPTMS-modified SiO₂, but not to bare SiO₂, indicating the chemical reactivity of epoxy groups on the modified nanoparticle with amines. Finally, fluorescently labeled bovine serum albumin (BSA) was used as a model protein to investigate the capacity of epoxy-SiO₂ nanoparticles for protein immobilization. The results showed that more proteins were immobilized on the particle with longer reaction time, higher NaCl concentration, lower pH, and less GPTMS content. More importantly, proteins bound to epoxy-SiO₂ nanoparticle were highly stable. Under optimized reaction conditions, as much as 25 mg BSA/g nanoparticle was covalently attached to the nanoparticle. The epoxy silane modification of silica nanoparticles offers a reactive surface for one-step and high-density protein immobilization. Supported by the National Basic Research Program of China (Grant No. 2006CB403303) and the National Natural Science Foundation of China (Grant Nos. 20621703, and 20825519 )