Rukshan T. Perera

Ion current rectification (ICR) refers to the asymmetric potential-dependent rate of the passage of solution ions through a nanopore, giving rise to electrical current-voltage characteristics that mimic those of a solid-state electrical diode. Since the discovery of ICR in quartz nanopipettes two decades ago, synthetic nanopores and nanochannels of various geometries, fabricated in membranes and on wafers, have been extensively investigated to understand fundamental aspects of ion transport in highly confined geometries. It is now generally accepted that ICR requires an asymmetric electrical double layer within the nanopore, producing an accumulation or depletion of charge-carrying ions at opposite voltage polarities. Our research groups have recently explored how the voltage-dependent ion distributions and ICR within nanopores can induce novel nanoscale flow phenomena that have applications in understanding ionics in porous materials used in energy storage devices, chemical sensing, and low-cost electrical pumping of fluids. In this Account, we review our most recent investigations on this topic, based on experiments using conical nanopores (10-300 nm tip opening) fabricated in thin glass, mica, and polymer membranes. Measurable fluid flow in nanopores can be induced either using external pressure forces, electrically via electroosmotic forces, or by a combination of these two forces. We demonstrate that pressure-driven flow can greatly alter the electrical properties of nanopores and, vice versa, that the nonlinear electrical properties of conical nanopores can impart novel and useful flow phenomena. Electroosmotic flow (EOF), which depends on the magnitude of the ion fluxes within the double layer of the nanopore, is strongly coupled to the accumulation/depletion of ions. Thus, the same underlying cause of ICR also leads to EOF rectification, i.e., unequal flows occurring for the same voltage but opposite polarities. EOF rectification can be used to electrically pump fluids with very precise control across membranes containing conical pores via the application of a symmetric sinusoidal voltage. The combination of pressure and asymmetric EOF can also provide a means to generate new nanopore electrical behaviors, including negative differential resistance (NDR), in which the current through a conical pore decreases with increasing driving force (applied voltage), similar to solid-state tunnel diodes. NDR results from a positive feedback mechanism between the ion distributions and EOF, yielding a true bistability in both fluid flow and electrical current at a critical applied voltage. Nanopore-based NDR is extremely sensitive to the surface charge near the nanopore opening, suggesting possible applications in chemical sensing.

An integrated current measurement system with ultra wide dynamic range is presented and fabricated in a 180-nm CMOS technology. Its dual-mode design provides concurrent voltage and frequency outputs, without requiring an external clock source. An integrator-differentiator core provides a voltage output with a noise floor of 11.6 fA/ [Formula: see text] and a -3 dB cutoff frequency of 1.4 MHz. It is merged with an asynchronous current-to-frequency converter, which generates an output frequency linearly proportional to the input current. Together, the voltage and frequency outputs yield a current measurement range of 155 dB, spanning from 204 fA (100 Hz) or 1.25 pA (10 kHz) to 11.6 μA. The proposed architecture's low noise, wide bandwidth, and wide dynamic range make it ideal for measurements of highly nonlinear electrochemical and electrophysiological systems.

Measured apparent activation energies, EA, of ion transport (K+ and Cl–) in conical glass nanopores are reported as a function of applied voltage (−0.5 to 0.5 V), pore size (20–2000 nm), and electrolyte concentration (0.1–50 mM). EA values for transport within an electrically charged conical glass nanopore differ from the bulk values due to the voltage and temperature-dependent distribution of the ions within the double layer. Remarkably, nanopores that display ion current rectification also display a large decrease in EA under accumulation mode conditions (at applied negative voltages versus an external ground) and a large increase in EA under depletion mode conditions (at positive voltages). Finite element simulations based on the Poisson–Nernst–Planck model semiquantitatively predict the measured temperature-dependent conductivity and dependence of EA on applied voltage. The results highlight the relationships between the distribution of ions with the nanopore, ionic current, and EA and their dependencies on pore size, temperature, ion concentration, and applied voltage.

The carcinogenic precursor benzo[a]pyrene (BP), a polycyclic aromatic hydrocarbon, is released into the environment through the incomplete combustion of hydrocarbons. Metabolism of BP in the human body yields a potent alkylating agent (benzo[a]pyrene diol epoxide, BPDE) that reacts with guanine (G) in DNA to form an adduct implicated in cancer initiation. We report that the α-hemolysin (αHL) nanopore platform can be used to detect a BPDE adduct to G in synthetic oligodeoxynucleotides. Translocation of a 41-mer poly-2'-deoxycytidine strand with a centrally located BPDE adduct to G through αHL in 1 M KCl produces a unique multi-level current signature allowing the adduct to be detected. This readily distinguishable current modulation was observed when the BPDE-adducted DNA strand translocated from either the 5' or 3' directions. This study suggests that BPDE adducts and other large aromatic biomarkers can be detected with αHL, presenting opportunities for the monitoring, quantification, and sequencing of mutagenic compounds from cellular DNA samples.

The identification and discrimination of four epigenetic modifications to cytosine in the proposed active demethylation cycle is demonstrated at the single-molecule level, without the need for chemical pretreatment or labeling. The wild-type protein nanopore α-hemolysin is used to capture individual DNA duplexes containing a single cytosine-cytosine mismatch. The mismatch is held at the latch constriction of α-hemolysin, which is used to monitor the kinetics of base-flipping at the mismatch site. Base-flipping and the subsequent interactions between the DNA and the protein are dramatically altered when one of the cytosine bases is replaced with methyl-, hydroxymethyl-, formyl-, or carboxylcytosine. As well as providing a route to single-molecule analysis of important epigenetic markers in DNA, our results provide important insights into how the introduction of biologically relevant, but poorly understood, modifications to cytosine affect the local conformational dynamics of a DNA duplex in a confined environment.