Quantitative measurement of nanoparticle uptake by flow cytometry illustrated by an interlaboratory comparison of the uptake of labelled polystyrene nanoparticles

Abstract Quantification of nanoparticle uptake into cells provides important information for both the assessment of novel nanomedicines and for nanosafety studies. Among several methods available to detect and/or quantify nanoparticle uptake, flow cytometry represents a robust high throughput method that allows measuring the internalisation of fluorescently labelled nanoparticles in thousands of individual cells in relatively short time. Several factors can influence and affect studies of nanoparticle uptake into cells, from the quality of the label and its stability, to the preparation of the nanoparticle dispersion, the way cells are exposed to the nanoparticles and several steps of sample preparation for flow cytometry measurement. Here we discuss the impact of all of these factors and methods to take them into account in order to avoid artefacts in the quantification of nanoparticle uptake and to ensure reproducibility. We then present a Standard Operating Procedure (SOP) for the quantification of nanoparticle uptake by flow cytometry, which has been developed within the European Research Infrastructure QualityNano by taking into account all of the described factors. Finally, we show the results obtained using the QualityNano SOP, demonstrating that with this SOP very good agreement in nanoparticle uptake measurements is achieved in independent laboratories by different operators using different instruments.