Playing the waiting game with aptamer probes

US nanotechnologists have solved a major problem hindering the development of highly sensitive and selective probes for use in complex biological fluids.   The team developed synthetic single-stranded oligonucleotides - called aptamers - as effective probes that could find use in a variety of medical, forensic and environmental applications.

Aptamers can be selected to bind any molecular target and can be produced rapidly in large quantities. They can also be attached to fluorescent compounds, which shine when aptamers bind to proteins of interest. 

The problem is that many biological compounds, including certain proteins and vitamins, naturally fluoresce when illuminated with ultraviolet light. This background fluorescence can mask the signal from an aptamer.

Weihong Tan from the University of Florida, Gainesville, has sidestepped the problem by attaching two fluorescent pyrene molecules to each end of an aptamer selected to bind with platelet-derived growth factor (PDGF). Raised levels of PDGF have been linked to several cancers, including breast and ovarian cancers. 

The aptamer changes shape when it binds with PDGF, bringing the two pyrene molecules together to form an excited dimer (excimer). The excimer fluoresces at a different wavelength to the individual pyrene molecules, and the fluorescence lasts much longer than that generated by most biological compounds (up to 100ns compared with less than 5ns).

Tan’s team found that, by waiting around 40ns after UV illumination, the fluorescence from the protein-bound aptamers was easily detectable because the background fluorescence had died down. The researchers are now adapting the technique to detect other protein biomarkers and are in discussions to develop a commercial detector for cancer biomarkers based on these probes. Jon Evans