Isotopically enriched nanoparticles in combination with mass spectrometry for the assessment of nanoparticle-biomolecule stoichiometries in engineered nanoassemblies

Highlights: • Direct stoichiometry determination between sulfur-containing NPs and biomolecules • Synthesis of engineered nanoparticles containing isotopically enriched sulfur • Analytical information relies on a simple accurate and precise sulfur isotope ratio. Determination of stoichiometries between engineered nanoparticles and species attached to their surface after chemical functionalization or interaction with biological media has been a hotspot during the last years, still unsatisfactory solved. Herein we propose a new approach only requiring for a simple mass spectrometric sulfur isotope ratio measurement in the final solution. Of course, enriched ³⁴S isotope has to be previously introduced in the engineered nanoparticle structure. Conversely to existing methodologies, mostly based on relative analyte measurements, proposed method does not require any previous knowledge of the molecule concentration in the starting solution. Such rather generic approach can be applied to study physical and/or chemical interactions between any sulfur-containing molecule and a wide variety of engineered nanoparticles containing sulfur as well, either in their own core-shell structure or in the ligand used for their surface functionalization (e.g. any thiol-derivative). In this paper, an inexpensive standard synthetic procedure was carried out to incorporate the required stable isotopic ³⁴S into the ZnS shell of CdSe quantum dots (QDs). Such isotopically enriched nanoparticles were then successfully applied to the quantitative assessment of stoichiometries between QDs and different target biomolecules, such as surface-functionalizing molecules (e.g. biotin) and proteins (e.g. bovine serum albumin). Of course, any biological conclusion and/or optimized bioconjugation conditions obtained using such isotopically enriched QDs analogues could be easily translated to nanoassemblies obtained using regular non-enriched QDs since they are chemically and physically identical.