Electrochemical Detection of Vascular Endothelial Growth Factor by an Aptamer-Based Bound/Free Separation System

Abstract

Vascular endothelial growth factor (VEGF) is expected to find application as a prognostic biomarker in cancer diagnosis. In this study, we measured VEGF using a simple bound/free separation system that utilizes an aptamer, whose capacity for hybridization with capture DNA immobilized on beads changes in the presence or absence of target molecules. Two systems were constructed using a VEGF aptamer selected by in vitro screening and a VEGF aptamer improved by a dimerization strategy. We can detect VEGF based on electrochemistry using both aptamers. We previously reported the detection of two proteins in addition to VEGF-IgE and thrombin-and the success of VEGF detection with this system suggests that this is a versatile system for the detection of various molecules.

Vascular Endothelial Growth Factor (VEGF) Detection Using an Aptamer and PNA-Based Bound/Free Separation System

“…VEGF detection systems have been commercialized and these kits can detect lower amount of VEGF than our assay. In addition, VEGF detection sensing systems using aptamers have been also reported [ 3 , 16 – 21 ]. Although an aptamer based B/F separation system using CaPNA is simple detection system, detection limit was not sufficient for the clinical applications.…”

“…We prepared CaPNA-immobilized beads by the same way as CaDNA [ 3 , 4 ]. We used NeutrAvidin beads with a near-neutral isoelectric point to reduce nonspecific binding of VEGF to the beads.…”

“…Using the complementary DNA of an aptamer, we can construct unique, versatile sensing systems. We have previously described an aptamer-based bound/free separation (B/F separation) system using complementary capture DNA (CaDNA) [ 3 , 4 ]. Target binding to the aptamer stabilizes the aptamer structure, which then inhibits the hybridization of complementary DNA to the aptamer.…”

Ultrasensitive SERS detection of VEGF based on a self-assembled Ag ornamented–AU pyramid superstructure

“…Other analytical platforms proposed for the quantification of VEGF include electrical detection of VEGF using Si nanowire field effect transistors (Lee et al, 2009), rolling circle amplification (Cheng et al, 2010a(Cheng et al, , 2010bSuzuki and Yokoyama, 2011), enhanced resonance light scattering (Chen et al 2012), aptasensor based on fluorescence polarization , fluorescent peptide conjugated nanopillar chip (Suzuki and Yokoyama, 2011), porosity induced hydrogel microspheres (Mohammad Al-Ameen, 2013) and optical and electrochemical sensors based on an aptasensor (Abe et al, 2012;Cho et al, 2012;Freeman et al, 2012). However, expensive instrumentation is involved in these methods.…”

Methods for Improving Aptamer Binding Affinity

“…SELEX involves simple separation steps of DNA or RNA that recognize target molecules from DNA or RNA libraries, and the repetition of these steps with a gradual increase in selection pressure leads to the isolation of aptamers with high affinity. Due to various advantages over antibodies, such as ease of chemical synthesis and modification, lower immunogenicity than antibodies, and the ability to refold after denaturation upon returning to the native condition, aptamers are versatile biomaterials that can be used as sensor elements, drugs, and drug delivery systems [3,4,5,6,7,8].…”