Interference-Free Detection of Genetic Biomarkers Using Synthetic Dipole-Facilitated Nanopore Dielectrophoresis

Abstract

The motion of polarizable particles in a non-uniform electric field, i.e., dielectrophoresis, has been extensively used for concentration, separation, sorting, and transport of biological particles, from cancer cells and viruses to biomolecules such as DNAs and proteins. However, current approaches to dielectrophoretic manipulation are not sensitive enough to selectively target individual molecular species. Here we describe the application of the dielectrophoretic principle for selective detection of DNA and RNA molecules using an engineered biological nanopore. The key element of our approach is a synthetic polycationic nanocarrier that selectively binds to the target biomolecules, dramatically increasing their dielectrophoretic response to the electric field gradient generated by the nanopore. The dielectrophoretic capture of the nanocarrier-target complexes is detected as a transient blockade of the nanopore ionic current while any non-target nucleic acids are repelled from the nanopore by electrophoresis and thus do not interfere with the signal produced by the target’s capture. Strikingly, we show that even modestly charged nanocarriers can be used to capture DNA or RNA molecules of any length or secondary structure and simultaneously detect several molecular targets. Such selective, multiplex molecular detection technology would be highly desirable for real-time analysis of complex clinical samples.

Single-molecule sensing of peptides and nucleic acids by engineered aerolysin nanopores

“…15). In a previous study of α-HL, K131D 7 , a mutant with a negatively charged ring at the trans entrance was able to capture the polycationic nanocarriers 29 . Here, we show that, as the positively charged amino acids in the cap region were replaced by the neutral amino acids, they are able to capture polycationic peptides.…”

A review of polystyrene bead manipulation by dielectrophoresis

“…The major issue is that individual biomolecules have very similar and very small induced dipole moments, and a high eld gradient is required to generate a substantial DEP force. 181 Thus, further research on the design of DEP systems is required for the effective uidic manipulation of multi-particles. Microfabrication techniques can help in achieving better designs, cost-effectiveness and efficiency.…”

Free-energy landscapes of membrane co-translocational protein unfolding

“…This closeness to the native state implies that at the transition state the oligonucleotide is threading the barrel and is consistent with the sensitivity of the refolding reaction to the applied voltage. This is relevant because the refolding opposing forces arising from the oligonucleotide re-entry 42 do not add up to rate limit the refolding reaction. Finally, the complete unfolding of the intermediate is irreversible because the unfolded polypeptide translocates through the pore (red, Fig.…”