This project focuses on discovering a new aptamer sequence that binds specifically and with high affinity to Brain Natriuretic Peptide (BNP), which is indicative of the early stages of heart failure. A library of random DNA sequences was generated synthetically, amplified by the polymerase chain reaction (PCR) and evaluated for specific binding to BNP using an in vitro selection technique known as systematic evolution of ligands by exponential enrichment (SELEX). The selective process was performed using BNP immobilized on an affinity chromatography column. Interacting aptamers were recovered, amplified by PCR using a 5’-phosphorylated reverse primer, and submitted to an enzymatic strand-separation reaction with lambda exonuclease before reiterating the process. After 10 and 15 SELEX cycles, selected aptamers were cloned, sequenced and their affinity and specificity with BNP investigated. Finally, the best aptamer was characterized by measuring its binding constant with BNP using surface plasmon resonance, and the structure of the complex is underway. In the future, this aptamer will be conjugated to nanoparticles and the resulting nanobiosensor used to devise a new detection method of BNP by surface enhanced resonance raman scattering, offering a top quality diagnosis and thus, allowing a faster initiation of appropriate treatment for heart failure.
Journal: TechConnect Briefs
Volume: 2, Technical Proceedings of the 2007 NSTI Nanotechnology Conference and Trade Show, Volume 2
Published: May 20, 2007
Pages: 287 - 290
Industry sectors: Advanced Materials & Manufacturing | Medical & Biotech
Topicss: Biomaterials, Materials Characterization & Imaging