Measurements of interactions between fluorescent molecules and polyethylene glycol self-assembled monolayers

Blocking non-specific binding of fluorescent biomolecules in single-molecule detection is one of the most important approach to minimize the background noise. Polyethylene glycol (PEG) and its derivatives are the most frequently-used self-assembled monolayers (SAMs) for surface passivation because they are particularly effective at reducing the adsorption of a majority of fluorescent biomolecules. Most studies about mechanism focus only on biomolecules themselves and sparse literature exists examining the interactions between commercial fluorophores and organosilane SAMs. The objective of this study was studying the interactions between fluorescein isothiocyanate (FITC) and PEG SAMs in solutions with different ionic strengths. Total internal reflection microscopy (TIRM) was utilized for quantitative analysis of the interactions. In low ionic strength solution, strong electrostatic attractions between PS-silica-FITC particles and PEG500 SAMs were detected even though both bodies were negatively charged. The origin of the attractive force was the charge correlations between the heterogeneous surfaces. In high ionic strength, the electrostatic attractions were completely screened, only left with the van der Waals attractions between the PS-silica-FITC particles and PEG SAMs. Due to the polarizability of FITC molecule, Hamaker constant was enlarged and the measured attractive forces depended on the thickness of PEG SAMs. The phenomenon can be explained by the mechanism of biomolecular resistance, including steric repulsion and hydration shell. The energy penalty of a small molecule is much less than that of a large biomolecule while penetrating into the PEG layers. This work reveals that the PEG SAMs not only fail to prevent the non-specific binding of FITC molecules, but also attract the fluorophores through van der Waals interactions.