Single-molecule detection represents the highest sensitivity in analytical techniques, enabling precise studies of individual molecular events. Methods based on single-molecule fluorescence energy transfer offer nanoscale distance measurements with significant implications for structural biology, biophysics, and material-biomolecule interfaces. Förster Resonance Energy Transfer (FRET) operates in the 3–10 nm range, typically involving two organic fluorophores as energy donor and acceptor. Graphene-based energy transfer (GET) extends this range to 10–40 nm, with a fluorophore as the energy donor and graphene acting as a broadband, unbleachable energy acceptor. FRET has been used predominantly to study protein conformational changes, but its measurements are often qualitative due to challenges in resolving nanometer-scale distances accurately. GET is a more straightforward quantitative method that enables precise z-localization of fluorophores. Notably, it has recently enabled real-time monitoring of DNA-protein interactions at the structural level [1]. However, graphene's hydrophobic nature and the strong influence of thickness on fluorescence lifetime measurements limit its biocompatibility to non-DNA systems and overall reproducibility.

Our research focuses on titanium carbide (Ti₃C₂Tₓ) MXene as a complementary, powerful energy transfer tool. MXenes possess unique advantages for studying biological assemblies that require hydrophilic surfaces, combining FRET-like short-distance sensitivity (1–10 nm) with GET's quantitative protocol, while offering unique material advantages such as hydrophilicity, biocompatibility, and thickness-independent energy transfer efficiencies.

This research line began by exploring MXene-DNA interactions with ensemble fluorescence spectroscopy measurements and molecular dynamics simulations. The interaction between MXene and DNA was found to be driven by salt bridges, coinciding with prior reports [2], with ssDNA and dsDNA lying horizontally on the surface while maintaining their native structure, approximately 1 and 1.3 nm away from the surface, respectively. In real-time hybridization experiments, we observed a fluorescence increase as ssDNA converted to dsDNA on the surface, suggesting that MXenes possess nanoscale sensitivity to sub-nanometer changes in the fluorophore position [3].

Next, we investigated MXenes’ fluorescence quenching mechanism and distance dependency of the energy transfer with single-molecule fluorescence microscopy and DNA origami nanopositioners using MXene thin films on glass. We positioned a single dye (ATTO 542) at defined distances from the surface (1–8 nm) via glycine-immobilized DNA origami nanostructures on MXene surfaces. Fluorescence quenching was observed between 1 to 8 nm, following a cubic distance dependence, consistent with the Förster mechanism observed in transparent conductors at the bulk level. These findings not only supported the hypothesis of MXenes' sub-nanometer sensitivity but also established them as hydrophilic, short-distance spectroscopic nanorulers uniquely positioned to operate in a distance regime inaccessible to GET. Additionally, the energy transfer efficiency was nearly independent of material thickness, enhancing robustness for sensing applications. 

Finally, we demonstrated the utility of MXene's sensitivity, robustness and hydrophilic, salt-driven interactions for leaflet-resolved, single-molecule biosensing of fluorescently-labeled, 5-nm thick supported lipid bilayers (SLBs) as model cell membranes, directly fused on the MXene surface [4]. Similar to typical SLBs formed on mica surfaces—where a thin hydration layer separates the bilayer from the surface to maintain biomimetic fluidity—MXenes also supported biomimetic SLBs, offering an advantage over other metallic surfaces and graphene. At the single-molecule level, we determined the SLB thickness by independently resolving each leaflet of the bilayer and the hydration layer separating it from the MXene surface. This capability to discern such fine structures highlights MXene substrates as uniquely suited to studying lipid-protein interactions, membrane organization, and dynamic hydration phenomena, potentially offering new insights into lipid biology and biomembrane research.