Quantification of Nanoparticles at the Single-cell Level: An Overview About State-of-the-art Techniques and Their Limitations

Dimitri Vanhecke; Laura Rodriguez-Lorenzo; Martin JD Clift; Fabian Blank; Alke Petri-Fink; Barbara Rothen-Rutishauser


Nanomedicine. 2014;9(12):1885-1900. 

In This Article


Until now, the research on NPs has been dedicated to a detailed description of the synthesis and characterization of NPs and their possible cellular reactions; however, very often the characterization of the biological part displays a great potential for improvement. The biomedical application of any NP requires a precise knowledge about the uptake mechanisms into any cell and the subsequent intracellular particle distribution. In addition, drug delivery of newly designed nanocarriers is one of the most promising fields in nanomedicine.

During recent years a large array of efficient and accurate techniques have been developed and optimized to detect NPs, locate the specific compartment and quantify the total number of intracellular NPs at the single-cell level. Up to now, no single technique exists that allows an optimal acquisition of intracellular NP quantities and their localization in specific compartments, each of the methods has unique strengths and weaknesses (for a summary please see Table 1).

The data gathered from many reports have demonstrated the usefulness of ICP techniques as an analytical tool for in vitro assays involving NPs. ICP allows for the efficient quantification of delivery parameters – that is, the differences in internalization over time, which would be difficult to examine by microscopic imaging techniques. Compared to image analysis, ICP is more sensitive to quantify small amounts of NPs within cells. ICP can sense differences of internalized gold concentrations at the order of one part per billion.[106]

Flow cytometry provides high throughput information coupled to differences in the cellular population. Combined with fluorescent biomarkers this method offers statistically sound data of the specific processes, such as NP uptake and/or association, by specific cell types in a diverse cellular landscape.

Microscopy techniques allow localization of NPs, such as determining if NPs are intracellular or only attached to the outer cell membrane. Furthermore, dynamic and kinetic informations in living (such as intracellular trafficking) is possible by light microscopy. For light, as well as electron, microscopy various quantification methods, as for example by means of stereology, have been developed. Unbiased stereology for quantitative image analysis is therefore an important tool, since qualitative or biased data can lead to misinterpretation, especially in the field of NP–cell interactions. By understanding the underlying mechanism of the NP–cell interface we will have the possibility to design and optimize new nanotechnology approaches from basic research concepts to medical applications.