New Fluorescent Nanoparticles for Ultrasensitive Detection of Nucleic Acids by Optical Methods

5 years ago

New Fluorescent Nanoparticles for Ultrasensitive Detection of Nucleic Acids by Optical Methods

Jørgen Kjems, Anders H. Okholm, Rasmus P. Thomsen, Kira Astakhova, Maria Taskova, Mads Westergaard Mulberg

For decades the detection of nucleic acids and their interactions at low abundances has been a challenging task that has thus far been solved by enzymatic target amplification. In this work we aimed at developing efficient tools for amplification-free nucleic acid detection, which resulted in the synthesis of new fluorescent nanoparticles. Here, the fluorescent nanoparticles were made by simple and inexpensive radical emulsion polymerization of butyl acrylate in the presence of fluorescent dyes and additional functionalization reagents. This provided ultra-bright macrofluorophores of 9–84 nm mean diameter, modified with additional alkyne and amino groups for bioconjugation. By using click and NHS chemistries, the new nanoparticles were attached to target-specific DNA probes that were used in fluorimetry and fluorescence microscopy. Overall, these fluorescent nanoparticles and their oligonucleotide derivatives have higher photostability, brighter fluorescence and hence dramatically lower limits of target detection than the individual organic dyes. These properties make them useful in approaches directed towards ultrasensitive detection of nucleic acids, in particular for imaging and in vitro diagnostics of DNA. Detection of low-abundance nucleic acids is a challenging task that has thus far been solved by enzymatic target amplification. Here we describe an alternative approach that combines organic fluorophore polymerization and site-specific bioconjugation for ultrasensitive detection of nucleic acids in vitro.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/cbic.201700125