Terminal protection of small molecule-linked ssDNA for label-free and highly sensitive colorimetric detection of folate receptor biomarkers

Abstract

Small molecule/protein interactions have a key role in drug discovery, clinic diagnosis and protein–metabolite interactions in biology. By using the specific interaction between folic acid (FA) and folate receptor (FR) as a model, the development of a label-free and sensitive colorimetric approach for the detection of the FR biomarker is described. The sensing approach relies on the coupling of the FR-induced terminal protection of FA-linked ssDNA strategy with significant signal amplification by self-assembled DNAzyme polymers. The FR binds to the FA-ssDNA and protects the FR/FA-ssDNA from digesting by exonuclease I. The terminal protected ssDNA further triggers autonomous self-assembly of two G-quadruplex sequence-containing hairpin DNAs into DNAzyme polymers, which result in intensified color change of the probe solution for label-free and highly sensitive colorimetric detection of FR. The terminal protection mechanism and the self-assembly formation of the DNAzyme polymers are characterized by using polyacrylamide gel electrophoresis, and the sensing parameters are optimized as well. Under optimal experimental conditions, the detection limit of 0.35 pM for FR can be obtained by using a UV-Vis spectrophotometer and the presence of as low as 5 pM of FR can be directly visualized by the naked eye. The developed method is also selective and can be applied to detect FR in serum samples, which makes this approach a sensitive platform for sensing different types of small molecule/protein interactions.