Quantum dot agglomerates in biological media and their characterization by asymmetrical flow field-flow fractionation

• AF4 was used to determine the precise size distribution of agglomerated QDs in media.
• Surface chemistry, media composition, and incubation time affect agglomeration status.
• Size distribution results correlate well with QD uptake in different cell lines.
• Cytotoxicity in certain cell lines was observed after exposure to agglomerated QDs.

The molecular composition of the biological environment of nanoparticles influences their physical properties and changes their pristine physicochemical identity. In order to understand, or predict, the interactions of cells with specific nanoparticles, it is critical to know their size, shape, and agglomeration state not only in their nascent state but also in biological media. Here, we use asymmetrical flow field-flow fractionation (AF4) with on-line multiangle light scattering (MALS), dynamic light scattering (DLS) and UV–Visible absorption detections to determine the relative concentration of isolated nanoparticles and agglomerates in the case of three types of semi-conductor quantum dots (QDs) dispersed in Dulbecco’s Modified Eagle Media (DMEM) containing 10% of fetal bovine serum (DMEM-FBS). AF4 analysis also yielded the size and size distribution of the agglomerates as a function of the time of QDs incubation in DMEM-FBS. The preferred modes of internalization of the QDs are assessed for three cell-types, N9 microglia, human hepatocellular carcinoma cells (HepG2) and human embryonic kidney cells (Hek293), by confocal fluorescence imaging of live cells, quantitative determination of the intracellular QD concentration, and flow cytometry. There is an excellent correlation between the agglomeration status of the three types of QDs in DMEM-FBS determined by AF4 analysis and their preferred mode of uptake by the three cell lines, which suggests that AF4 yields an accurate description of the nanoparticles as they encounter cells and advocates its use as a means to characterize particles under evaluation.

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