Carbon Dot-Based Fluorosensor Shows Superior Performance in Enterovirus RNA Detection

Scientists from the University of Jyväskylä's Nanoscience Center in Finland have introduced a novel, label-free ratiometric fluorosensor intended for the sensitive and specific detection of enteroviral RNA. The study aims to provide even more sophisticated and efficient detection techniques, highlighting the value of interdisciplinary cooperation in tackling global health issues.

Recent pandemics have shown that viruses are a serious threat to world health. Preventing new outbreaks requires early identification and detection. Despite their effectiveness, traditional detection techniques often fail to provide spatiotemporal information about the release of the virus genome.

This interdisciplinary effort, combining expertise from biology, chemistry, and physics, marks a significant advancement in viral detection technology. We have developed an enhanced ratiometric fluorosensor using carbon dots (CDs) functionalized with Probe (single-stranded complementary oligonucleotide fragment) and ethidium bromide (EB), for detection of enteroviral RNA.”

Jussi Toppari, Professor, School of f Physics, University of Jyväskylä

Innovative Ratiometric Fluorosensor for Viral Detection

Due to their easy synthesis, remarkable photostability, adjustable photoluminescence, outstanding aqueous solubility, biocompatibility, and adaptable surface functionalities for ligand conjugation, fluorescent nanoparticles have become extremely effective tools for bioanalyte sensing.

CDs are at the forefront of this field. Due to these special qualities, CDs are positioned to revolutionize the biosensing industry.

This so-called Functionalized Sensor (Func Sensor), where CDs are functionalized i.e., covalently bonded with the probe clearly outperforms the more traditional approach of Non-Functionalized Sensor (Non-Func Sensor) which is a simple mixture of CDs, probe, and EB.”

 Amar Raj, Doctoral Researcher, University of Jyväskylä

The presence of target DNA, which hybridizes with the probe in both sensors, increases EB fluorescence, whereas electron transfer causes a slight change in CD fluorescence, allowing for ratiometric detection and making the sensors ultrasensitive.

The Non-Func Sensor showed a lower sensitivity with target DNA and was not effective with real enteroviral RNA samples, while the Func Sensor showed a higher sensitivity with DNA and real viral RNA, exhibiting clearly improved selectivity.”

Abhishek Pathak, Postdoctoral Researcher, University of Jyväskylä

The enhanced charge transfer brought about by covalent functionalization is responsible for the Func Sensor's superior performance.

“Our proof-of-principle study highlights the importance of covalent immobilization of the probe for improved electron transfer between CDs and EB and thus enhanced performance and demonstrates the suitability of the Func sensor for practical applications in rapid, real-time and precise in situ detection of viral RNA,” said Varpu Marjomäki, Professor of Cell and Molecular Biology from the University of Jyväskylä. 

In particular, the study demonstrates that the Func sensor can identify enteroviral RNA release from the capsid in vitro in real-time.

“This means that the Func sensor can be used as a novel viral RNA sensing platform which offers a much-needed possibility to detect real-time viral RNA appearance during infection,” said Marjomäki.  

Towards Safer Research 

The research team's groundbreaking work clarifies the charge transfer mechanisms between fluorophores in addition to demonstrating a novel technique for detecting viral RNA. Building on this achievement, the research team is currently attempting to strengthen the system by substituting the much safer, less cytotoxic, or biocompatible dyes for the potentially dangerous dye ethidium bromide.

“This enhancement will further improve the safety and efficacy of in vivo viral RNA detection,” explained Pathak.