Abstract
Fluorescence is a powerful tool for mapping biological events in real-time with high spatial resolution. Ultra-bright probes are needed in order to achieve high sensitivity: these probes are typically obtained by gathering a huge number of fluorophores in a single nanoparticle (NP). Unfortunately this assembly produces quenching of the fluorescence because of short-range intermolecular interactions. Here we demonstrate that rational structural modification of a well-known molecular fluorophore N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) (NBD) produces fluorophores that self-assemble in nanoparticles in the biocompatible environment without any dramatic decrease of the fluorescence quantum yield. Most importantly, the resulting NP show, in an aqueous environment, a brightness which is more than six orders of magnitude higher than the molecular component in the organic solvent. Moreover, the NP are prepared by nanoprecipitation and they are stabilized only via non-covalent interaction, they are surprisingly stable and can be observed as individual bright spots freely diffusing in solution at a concentration as low as 1 nM. The suitability of the NP as biocompatible fluorescent probes was demonstrated in the case of HeLa cells by fluorescence confocal microscopy and MTS assays.
Highlights
We describe the synthesis and the properties of a new family of molecular fluorophores (MF), which are water insoluble and do not suffer from aggregation caused quenching (ACQ), and we demonstrate that a rational design of the molecular structure allows us to achieve a molecular unit that self-organizes in highly bright, and fluorescent NPs which are very stable in phosphate-buffered saline (PBS) solution
Summarizing the results so far discussed, we found that molecule 1 does not form stable NP upon nanoprecipitation in our experimental conditions
The brightness of 3NP is about two orders of magnitude lower than the one of 2NP. Such a difference is so relevant that while 2NP can be clearly tracked at the single NP level in solution at very low concentration by fluorescence microscopy, 3NP cannot be observed with the same technique
Summary
Fluorescence imaging is a not invasive, highly sensitive, technique that allows to investigate biological organisms with high tridimensional resolution in real time, by making use of suitable fluorescent contrast agents (Rio-Echevarria et al, 2011; Cauzzi et al, 2012; Chen et al, 2015; Grimm et al, 2015; Lee et al, 2015; Mei et al, 2015; Tang et al, 2015; Antaris et al, 2016; Proetto et al, 2016; Xu et al, 2016). By comparing the NP to the molecular precursors, an increase of the brightness of about five order of magnitude, with respect to the fluorophore in organic solvent, could be estimated as a result of the self-assembly.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
