Studying aptamer binding to cancer cell membrane receptors by fluorescence correlation spectroscopy.

摘要

Aptamers are short strands of oligonucleotides, designed to have excellent binding affinity and selectivity towards certain target molecules. Due to their capacity to bind tightly and specifically to their ligands, aptamers have applications in fields ranging from medicine to biochemical research. Fluorescence correlation spectroscopy (FCS) is being recognized as a highly sensitive method for the investigation of molecular interactions. By using this technique, association and dissociation rate constants, and binding constants can be obtained from the analysis of the Brownian motion of fluorescently labeled probes. The present study is focused on cancer cell-specific aptamers binding to their target. For determination of the dissociation rate constant, a binding curve is obtained by incubating the target cells with different concentrations of fluorescently-labeled aptamer. The binding constants are then compared to the ones that have been reported by flow cytometry. Further, a method for calculation of the aptamer receptor density on the cell membrane is proposed. The study will show that FCS is a reliable method for the determination of binding properties of aptamer-target complexes even when used in vivo.