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Measuring stepwise binding of a thermally fluctuating particle to a cell membrane without labeling
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Researchers used optical trapping and interferometry to detect individual binding events between polystyrene beads and macrophage cell membranes without fluorescent labeling, tracking nanoscale position and orientation changes. The technique enables highly sensitive study of how micro- and nanoparticles interact with cell surfaces, relevant to understanding microplastic uptake by immune cells.
ABSTRACT Thermal motions enable a particle to probe the optimal interaction state when binding to a cell membrane. However, especially on the scale of microseconds and nanometers, position and orientation fluctuations are difficult to observe with common measurement technologies. Here we show that it is possible to detect single binding events of IgG-coated polystyrene beads, which are held in an optical trap nearby the cell membrane of a macrophage. Changes in the spatial and temporal thermal fluctuations of the particle were measured interferometrically and no fluorophore labelling was required. We demonstrate both by Brownian dynamic simulations and by experiments that sequential step-wise increases in the force constant of the bond between a bead and a cell of typically 20 pN / µm are clearly detectable. In addition, this technique provides estimates about binding rates and diffusion constants of membrane receptors. The simple approach of thermal noise tracking points out new strategies in understanding interactions between cells and particles, which are relevant for a large variety of processes including phagocytosis, drug delivery or the effects of small microplastics and particulates on cells. SIGNIFICANCE Interactions of cells with nearby particles, e.g. bacteria, viruses or synthetic material, is a very fundamental and complex process, often deciding about the cellular fate. The investigation of binding processes between particle and cell is typically investigated by fluorescence techniques, where fluorophores often hinder the molecular interaction of specific binding partners. Therefore, label-free detection or imaging techniques are essential, which are hardly available especially for live cell investigations. Molecular binding is based on thermal position and orientation fluctuations of the binding partners to find the best interaction state. Here, we present a label-free measurement technique that allows us to detect multiple stepwise binding events of molecules on an optically trapped particle close to the cell membrane.
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