Tunable resistive pulse sensing (TRPS) is a technology capable of characterising the size, concentration and charge of nanoparticles in solution. By applying a combination of pressure and voltage, sample particles are driven through a nanopore under an applied electric field. As analytes pass through the nanopore, there is a resulting drop in current. The change in current (referred to as a single ‘blockade’ event) is recorded and can be analysed to determine the physical characteristics of the sample (concentration) and individual particles (size and zeta potential). TRPS has been used to characterise exosomes and other extracellular vesicles, as well as monoclonal antibody preparations and vaccine candidates.
1. Zeta potential of EVs from CRISPR-edited cells
TRPS was used as part of a study aimed at understanding the renal trafficking of renally-derived EVs. Disruptions to single genes Pkd1 and Pkd2 (which code for proteins Polycystin 1 and 2, respectively), introduced using CRISPR/CAS9, had a significant impact on the zeta potential of EVs produced by those cells. Compared to EVs from Pkd1-disrupted cells, EVs from Pkd2-disrupted cell lines had a more negative zeta potential.1
2. Size profiling of EVs from human corneal stromal stem cells
TRPS was used to identify the size distribution of EVs from corneal stromal stem cells, which are being investigated for their therapeutic potential following corneal stromal injury. Anti-scarring effects, associated with miR-29a expression, were validated in vitro and in mouse model studies.2
3. Characterisation of clinically relevant neutrophil-derived EVs
Bonifay et al. (2022) describe a new strategy for the measurement of circulating neutrophil-derived EVs, which have been reported to be elevated in a variety of infectious and cardiovascular diseases. As part of this development, TRPS was used to confirm that that vesicle size in the preparations were in accordance with that of EVs.3
4. EVs derived from human Sertoli cells
TRPS was among the characterisation methods used in an investigation of EVs isolated from human Sertoli cells. The study aimed to characterise protein and miRNA cargo in EVs, laying the foundation for further studies of Sertoli cell-derived EVs and their potential roles in spermatogenesis and male infertility.4
5. Exploring the functionality of the EV protein corona
In a recent study by Wolf et al. (2022), TRPS was used to measure zeta potential of EVs in the absence or presence of a re-established protein corona. Here, a more positive zeta potential was observed in ultracentrifuged (identified as ‘corona-depleted’) EVs compared to when the protein corona was artificially re-established. Altogether, this work highlights the ongoing need to determine the relative contribution of cells, EVs and soluble factors in relevant therapies under development, and raises questions about the EV protein corona and implications for the development of clinical applications.5
