Patch-clamp Amplifier Research Articles

Shedding Light on Cardiac Excitation: In Vitro and In Silico Analysis of Native Ca2+ Channel Activation in Guinea Pig Cardiomyocytes Using Organic Photovoltaic Devices.

This study aims to explore the potential of organic electrolytic photocapacitors (OEPCs), an innovative photovoltaic device, in mediating the activation of native voltage-gated Cav1.2 channels (ICa,L) in Guinea pig ventricular cardiomyocytes. Whole-cell patch-clamp recordings were employed to examine light-triggered OEPC mediated ICa,L activation, integrating the channel's kinetic properties into a multicompartment cell model to take intracellular ion concentrations into account. A multidomain model was additionally incorporated to evaluate effects of OEPC-mediated stimulation. The final model combines external stimulation, multicompartmental cell simulation, and a patch-clamp amplifier equivalent circuit to assess the impact on achievable intracellular voltage changes. Light pulses activated ICa,L, with amplitudes similar to voltage-clamp activation and high sensitivity to the L-type Ca2+ channel blocker, nifedipine. Light-triggered ICa,L inactivation exhibited kinetic parameters comparable to voltage-induced inactivation. OEPC-mediated activation of ICa,L demonstrates their potential for nongenetic optical modulation of cellular physiology potentially paving the way for the development of innovative therapies in cardiovascular health. The integrated model proves the light-mediated activation of ICa,L and advances the understanding of the interplay between the patch-clamp amplifier and external stimulation devices. Treating cardiac conduction disorders by minimal-invasive means without genetic modifications could advance therapeutic approaches increasing patients' quality of life compared with conventional methods employing electronic devices.

Long-term memory in Staphylococcus aureus α-hemolysin ion channel kinetics.

The kinetics of an ion channel are classically understood as a random process. However, studies have shown that in complex ion channels, formed by multiple subunits, this process can be deterministic, presenting long-term memory. Staphylococcus aureus α-hemolysin (α-HL) is a toxin that acts as the major factor in Staphylococcus aureus virulence. α-HL is a water-soluble protein capable of forming ion channels into lipid bilayers, by insertion of an amphipathic β-barrel. Here, the α-HL was used as an experimental model to study memory in ion channel kinetics. We applied the approximate entropy (ApEn) approach to analyze randomness and the Detrended Fluctuation Analysis (DFA) to investigate the existence of long memory in α-HL channel kinetics. Single-channel currents were measured through experiments with α-HL channels incorporated in planar lipid bilayers. All experiments were carried out under the following conditions: 1 M NaCl solution, pH 4.5; transmembrane potential of + 40mV and temperature 25 ± 1°C. Single-channel currents were recorded in real-time in the memory of a microcomputer coupled to an A/D converter and a patch-clamp amplifier. The conductance value of the α-HL channels was 0.82 ± 0.0025 nS (n = 128). The DFA analysis showed that the kinetics of α-HL channels presents long-term memory ([Formula: see text] = 0.63 ± 0.04). The ApEn outcomes showed low complexity to dwell times when open (ApEno = 0.5514 ± 0.28) and closed (ApEnc = 0.1145 ± 0.08), corroborating the results of the DFA method.

A 146-dB-Ohm Gain 14-pARMS Noise Patch-Clamp Amplifier for Whole-Cell Ion Current Detection

The membrane clamp technique is an important tool to reflect the electrophysiological characteristics of cells by recording the ionic currents of cellular channels. Embedded systems work in systems designed for specific user groups and are used to implement specific functions. The membrane clamp amplifier built with embedded technology has the advantages of miniaturization, specialization, low power consumption, high integration, high resource utilization, and a long life cycle, which can avoid the inconvenience to developers and experimenters due to the update of general-purpose computer software and hardware. In this paper, a patch-clamp amplifier (PCA) based on a transimpedance amplifier (TIA) is proposed, which includes a glass microelectrode series resistance/capacitance compensation circuit and feedback resistor parasitic capacitance compensation. The prototype is designed using 180 nm CMOS technology and occupies an area of 720 μ m × 630 μ m . The prototype achieves a transimpedance gain of 145.6 dB-Ohm, a -3 dB bandwidth greater than 15 kHz, and an input reference noise current of 13.98 pARMS. Experimental results show that the proposed PCA is capable of compensating up to 30 pF of electrode capacitance and up to 10 MΩ for 86% of the series resistance, respectively. In addition, when the feedback resistor parasitic capacitance compensation circuit is enabled, the overshoot phenomenon disappears, overcoming the shortcomings of the conventional diaphragm clamp amplifier current clamp and maintaining the original key performance.

Kinetic and thermodynamic modeling of a voltage-gated sodium channel.

Like all biological and chemical reactions, ion channel kinetics are highly sensitive to changes in temperature. Therefore, it is prudent to investigate channel dynamics at physiological temperatures. However, most ion channel investigations are performed at room temperature due to practical considerations, such as recording stability and technical limitations. This problem is especially severe for the fast voltage-gated sodium channel, whose activation kinetics are faster than the time constant of the standard patch-clamp amplifier at physiological temperatures. Thus, biologically detailed simulations of the action potential generation evenly scale the kinetic models of voltage-gated channels acquired at room temperature. To quantitatively study voltage-gated sodium channels' temperature sensitivity, we recorded sodium currents from nucleated patches extracted from the rat's layer five neocortical pyramidal neurons at several temperatures from 13.5 to 30°C. We use these recordings to model the kinetics of the voltage-gated sodium channel as a function of temperature. We show that the temperature dependence of activation differs from that of inactivation. Furthermore, the data indicate that the sustained current has a different temperature dependence than the fast current. Our kinetic and thermodynamic analysis of the current provided a numerical model spanning the entire temperature range. This model reproduced vital features of channel activation and inactivation. Furthermore, the model also reproduced action potential dependence on temperature. Thus, we provide an essential building block for the generation of biologically detailed models of cortical neurons.

Ion Channel-Based Reporters for cAMP Detection.

In the last 20years tremendous progress has been made in the development of single cell cAMP sensors. Sensors are based upon cAMP binding proteins that have been modified to transduce cAMP concentrations into electrical or fluorescent readouts that can be readily detected using patch clamp amplifiers, photomultiplier tubes, or cameras. Here, we describe two complementary approaches for the detection and measurement of cAMP signals near the plasma membrane of cells using cyclic nucleotide (CNG) channel-based probes. These probes take advantage of the ability of CNG channels to transduce small changes in cAMP concentration into ionic flux through channel pores that can be readily detected by measuring Ca2+ and/or Mn2+ influx or by measuring ionic currents.

Rosuvastatin Reduces L-Type Ca2+ Current and Alters Contractile Function in Cardiac Myocytes via Modulation of β-Adrenergic Receptor Signaling.

Rosuvastatin is one of the most used statins to lower plasma cholesterol levels. Although previous studies have reported remarkable cardiovascular effects of rosuvastatin (RSV), the mechanisms of these effects are largely unknown. In this study, we investigated the acute effects of RSV on L-type Ca2+ currents and contractile function of ventricular myocytes under basal conditions and during β-adrenergic stimulation. The effects of RSV were investigated in freshly isolated adult rat ventricular myocytes. L-type Ca+2 currents and myocyte contractility were recorded using patch-clamp amplifier and sarcomere length detection system. All experimental recordings were performed at 36 ± 1°C. L-type Ca+2 currents were significantly reduced with the administration of 1μM RSV (~ 24%) and this reduction in Ca2+ currents was observed at almost all potential ranges applied. Suppression of L-type Ca2+ current by RSV was prevented by adenylyl cyclase (AC) and protein kinase A (PKA) inhibitors SQ 22536 and KT5720, respectively. However, inhibition of Rho-associated kinases (ROCKs) by Y-27632 or nitric oxide synthase (NOS) by L-NAME failed to circumvent the inhibitory effect of RSV. Finally, we examined the effect of RSV during β-adrenergic receptor stimulation by isoproterenol and observed that RSV significantly suppresses the β-adrenergic responses in both L-type Ca2+ currents and contraction parameters. In conclusion, RSV modulates the β-adrenergic signaling cascade and thereby mimics the impact of β-adrenergic receptor blockers in adult ventricular myocytes through modulation of the AC-cAMP-PKA pathway.

Comparison of Deep Learning Models for Fully Automated Single Channel Idealization

Single ion channel analysis can reveal high resolution data about kinetics of single molecules, but has been described as “slow and laborious”. Typically, the first stage in such analysis is the creation of an idealized record with each timepoint classified as the number ion channels open. Whilst there have been a number of semi-automatic methods developed to assist with this, in situations with multiple channels in the patch or generally poor signal to noise ratio, these methods could be difficult to upscale these analysis for industrial use. With this in mind we are working on artificial intelligence methods that may one day supersede existing methods. One recent approach, called DeepChannel, used developments in deep learning (a branch of artificial intelligence) to fully idealize data fully automatically. Deep learning is heavily dependent on large datasets to train, which in DeepChannel's case were generated by passing simulated ion channel events through a real patch clamp amplifier. This method was thus limited to real-time recording resulting in the size of datasets to still be relatively small. Here, we describe new, completely digital data generation pipeline simulated data ion channel data and use these to train and compare new and old deep learning models including one based on the acclaimed image classification model “ResNet”. Additionally, we then use these models to idealise real world data, and compare with an expert annotation to judge relative model performance. Deep learning models show great potential for fully automated single molecule analysis and in the future we will release example models as a web app for easy use by other ion channel biologists.

Dynamic Clamp on a Windows PC.

Dynamic clamp is a powerful tool for interfacing computational models and real cells. We describe here how to set up and carry out dynamic clamp experiments using a patch clamp amplifier, a National Instruments data acquisition card, and the freely available QuB software that operates on a PC running MS Windows.

Patch Clamp Technology in the Twenty-First Century.

In the almost four decades since its inception, the patch clamp technique has transitioned from a specialist skill to a method commonly used among many others in a lab. Development of patch clamp instrumentation has not been steady: A boost of product releases in rapid succession by multiple manufacturers in the 1990s had slowed to a trickle by the mid-2000s. In 2016, Sutter Instrument's entry into the market of turnkey patch clamp amplifier systems, defined as an amplifier with matching data acquisition hardware and software, caused a fresh breeze in a field in danger of going stale. Sutter has meanwhile completed the product line, culminating in the flagship dPatch® Ultra-fast, Low-noise Digital Amplifier. The dPatch System constitutes a contemporary, digital design that features many firsts, including digital signalcompensation, an extremely high bandwidth and fully integrated dynamic clamp capability, paired with the increasingly popular SutterPatch® Software.This chapter compares feature sets of the new Sutter instrumentation with the established platforms by the other two providers of turnkey systems, Axon Instruments by Molecular Devices and HEKA Elektronik by Harvard Bioscience. A variety of products from other manufacturers, who rely on combination with components from other sources rather than offering turnkey systems, are listed, but for their conceptual diversity not compared at a great level of detail. The chapter further covers architectural considerations for patch clamp systems, headstage design, data acquisition strategies and efficient structuring of the recorded data, controlling and monitoring periphery, advanced technologies, such as software lock-in amplifier capability and dynamic clamp features, and application modules for efficient analysis of action potentials and postsynaptic events.

Signal Processing for Single Biomolecule Identification Using Nanopores: A Review

Nanopore sensors provide an innovative platform for rapid and label-free biomolecule detection. The analytical information extracted from resistive pulse sensing (RPS) measurement, though powerful has several limitations. To address these challenges, various signal conditioning algorithms have been researched which include efficient clustering and classification of data, applying probabilistic models to reduce information overlap and also leveraging transient analysis of ionic current instead of the steady state RPS measurement. Additionally, as the current is typically in few picoampere regime with high transition rates, its faithful measurement requires precision instrumentation like a patch clamp amplifier with low noise and high bandwidth. However, for practical deployment of such systems, it is necessary to develop portable electronic interface. Significant efforts have been directed globally to develop on-chip customized integrated circuits with low noise, high bandwidth amplifiers. Thus, it is absolutely timely and pertinent to conduct a review on the various signal processing schemes and electronic interfaces for single biomolecule identification using nanopores which will enable to comprehend the current status for its deployment as a commercial platform for genomics and ultrasensitive detection. To the best of our knowledge, the data extraction methodologies and electronic interface schemes for nanopores have not yet been summarized. In this paper, a comprehensive review emphasizing the signal processing algorithms for DNA sequencing and protein identification in complex analyte using functionalized and non-functionalized nanopores has been presented along with the on-chip low noise electronic interfaces required for field deployable measurement.

Transport Properties of Gramicidin A Ion Channel in a Free-Standing Lipid Bilayer Filled With Oil Inclusions.

Ion channels are key proteins in mammalian cell membranes. They have a central role in the physiology of excitable cells such as neurons, muscle, and heart cells. They also play a crucial role in kidney physiology. The gramicidin ion channel is one of the most studied ion channels, in particular it was intensively employed to investigate the lipid–protein interactions in model cell membranes. For example, even though the sequence of gramicidin is extremely hydrophobic, its motion is impaired in membrane bilayer, i.e., it does not rapidly flip to the other membrane leaflet, and low channel activity were observed when gramicidin is added asymmetrically to only one leaflet of a model cell membrane. In this article, we study the transport properties of gramicidin channel in a heterogeneous model membrane. Using microfluidics, we are forming freestanding bilayers as model cell membranes including heterogeneous domains that are created by oil inclusions. The presence of oil inclusions is then demonstrated by measuring the bilayer capacity via a patch-clamp amplifier and fluorescent confocal inspection. Based on electrophysiological and optical measurements Gramicidin A (gA) ion channels are dispersed into the buffer phases on both side of the formed lipid bilayer and insert spontaneously into the bilayer upon formation. The presence of functional Gramicidin A is then demonstrated by measuring conductivity signals. Based on electrophysiological and optical measurements, we explore the consequence of the presence of these oil inclusions on the functionality of incorporated gA ion channels. For low oil concentration, we measure a decrease of gA transport properties due to the reduction of the bilayer tension. For large oil concentration, we measure a saturation of gA transport properties due to an increase of the bilayer thickness.

Analysis of Membrane Protein Deinsertion-Associated Currents with Nanoneedle-Supported Bilayers to Discover Pore Formation Mechanisms.

Analysis of the pore formation mechanisms of biological nanopores can provide insight into pore-forming peptide-induced diseases and into the characterization of nanopores employed in sensing methods. Evaluation of pore formation mechanisms is typically performed using microscopy including atomic force microscopy, transmission electron microscopy, as well as electrically via channel current measurements using a patch-clamp amplifier. However, due to the relatively low temporal resolution of the above-mentioned microscopy techniques and the low analysis accuracy of the channel current measurements, new analytical methods are required. Here, we describe a new analytical strategy to measure and analyze both ionic currents associated with biological nanopore insertion and deinsertion into and out of lipid bilayers to determine pore formation mechanisms for several representative proteins. The current changes associated with protein deinsertion are monitored as the lipid membrane leaflets are pulled apart-a unique phenomenon enabled by our gold nanoneedle measurement probe. This deinsertion current analysis (DiCA) is performed using a gold nanoneedle-supported lipid bilayer at which a bilayer membrane is formed by bringing together two lipid monolayers on the surface of the nanoneedle and at the interface of an aqueous solution and a lipid/oil mixture. The lipid bilayer can be pulled apart by removing the nanoneedle from this interface. In this study, we demonstrate the determination of pore formation mechanisms for four different pore-forming proteins and peptides-α-hemolysin, streptolysin O, alamethicin, and amyloid β 1-42 using DiCA. As a result, we successfully discern the pore formation mechanism, either addition or expansion, for each protein/peptide by analyzing the ratio and magnitude of insertion and deinsertion current events.

Developing a Microfabricated Lab-On-A-Chip Device for Patch-Clamp Applications with Internal Solution Exchange

Patch-clamp electrophysiology is a widely utilized technique in both academic and industrial laboratories to assess the function of ion channels. Despite its importance, conventional patch-clamp experiments remain inherently difficult to perform, laborious, and suffer from low data throughput. Automated patch-clamp systems have since been developed to address these issues, but have done so to the detriment of the versatility and affordability of the technique. Here, we have developed a new chip-based patch-clamp system, dubbed the SMPL system, that has been specifically microfabricated to deliver high quality patch-clamp recordings while improving upon the versatility of other chip-based patch-clamp systems. The SMPL chip is compatible with any adherent cell type, and can be used to culture and assess the electrophysiological properties of induced pluripotent stem cell (iPSC)-derived and primary cultures in addition to more conventional, stable-expression cell lines. The chip contains four independent microfluidic channels, each linked to a micropatterned patch-site, that enables for up to four sequential recordings to be taken. These microchannels also serve to allow intracellular solution to be exchanged during the course of an experiment. Since the chip has been fabricated entirely of optically transparent materials, imaging experiments can also be performed in parallel to electrophysiological assessments. The SPML chip connects to any existing patch-clamp amplifier using an affordable universal adapter and switchbox, allowing the user to decide which of the four sites is to be accessed. We are currently evaluating the capabilities of this technology applied to recording lipid bilayers and ion channels reconstituted in lipid vesicles.

Development of Solid-State Nanopore Life Detection Technology

Biomarkers collected from past and present life on Earth are a central guide in the development of flight-ready technologies designed to search for life on other planets and moons. Data from numerous laboratory and field studies have revealed the importance of incorporating capabilities that minimize the risk of false negatives or false positives during life detection missions. Biomolecules are the most unambiguous and information-rich of all known biosignatures. Indeed, the identification in a sample of biopolymers akin to DNA, RNA or proteins, would be difficult to refute as a successful life detection experiment. In the case of Mars, the search for biomolecules is inescapable, but few technological solutions exist for in situ identification. Thus, our work has focused on the development of technologies designed to detect biochemical polyelectrolyte molecules, which have been argued to be a universal signature for life due to their ability to store information while reducing their tendency to adopt complex tertiary structures that would prohibit their replication. To that end we have developed a solid-state nanopore device that has the ability to detect electrically charged organic polymers, such as those found in deoxyribonucleic acid. Our nanopore technology, when used as a biosensor, consists of two or three ion-filled chambers separated by voltage-biased thin layer (Figure 1). Nanopores are milled into the thin layer, allowing the chambers to be connected electrically via an ionic solution. Polyelectrolytes are introduced into one of the chambers, and a patch clamp amplifier is connected to the chip via electrodes that serve as a voltage source and an ammeter. When a voltage is applied across the membrane, the ionic current going only through this pore is measured. The polyelectrolytes are captured by the electric field in the pore, then and electrophoretically drawn through the nanopore (translocation) causing the nanopore conductance to decrease. This conductance decrease, or transient current blockage, is evaluated for: 1) mean blockage current (which is caused by the physical passage of the polyelectrolyte through the nanopore, and is measured by a sensitive ammeter in pA); 2) translocation duration (physical time of the translocation, for DNA this is typically 30 bases per second when translocation is controlled by a molecular motor [1]); and 3) integrated area of a blockage, revealing key features of the particle. Solid-state nanopore technology is being aggressively advanced for use as an electrochemistry sensor for a wide variety of biological molecules. Testing of the solid-state nanopore membranes includes the establishment of criteria used to distinguish informational from non-informational output signals. Our work highlights past, current, and future efforts in the development of solid-state nanopore technology for life detection.

Precise Construction and Tuning of an Aerolysin Single-Biomolecule Interface for Single-Molecule Sensing

Precise Construction and Tuning of an Aerolysin Single-Biomolecule Interface for Single-Molecule Sensing

Simultaneous recording of electrical activity and the underlying ionic currents in NG108-15 cells cultured on gold substrate

This paper shows the simultaneous recording of electrical activity and the underlying ionic currents by using a gold substrate to culture NG108-15 cells. Cells grown on two different substrates (plastic Petri dishes and gold substrates) were characterized quantitatively through scanning electron microscopy (SEM) as well as qualitatively by optical and atomic force microscopy (AFM). No significant differences were observed between the surface area of cells cultured on gold substrates and Petri dishes, as indicated by measurements performed on SEM images. We also evaluated the electrophysiological compatibility of the cells through standard patch-clamp experiments by analyzing features such as the resting potential, membrane resistance, ionic currents, etc. Cells grown on both substrates showed no significant differences in their dependency on voltage, as well as in the magnitude of the Na+ and K+ current density; however, cells cultured on the gold substrate showed a lower membrane capacitance when compared to those grown on Petri dishes. By using two separate patch-clamp amplifiers, we were able to record the membrane current with the conventional patch-clamp technique and through the gold substrate simultaneously. Furthermore, the proposed technique allowed us to obtain simultaneous recordings of the electrical activity (such as action potentials firing) and the underlying membrane ionic currents. The excellent conductivity of gold makes it possible to overcome important difficulties found in conventional electrophysiological experiments such as those presented by the resistance of the electrolytic bath solution. We conclude that the technique here presented constitutes a solution to the problem of the simultaneous recording of electrical activity and the underlying ionic currents, which for decades, had been solved only partially.

Artifactual hyperpolarization during extracellular electrical stimulation: Proposed mechanism of high-rate neuromodulation disproved

BackgroundKilohertz-frequency electric field stimulation (kEFS) applied to the spinal cord can reduce chronic pain without causing the buzzing sensation (paresthesia) associated with activation of dorsal column fibers. This suggests that high-rate spinal cord stimulation (SCS) has a mode of action distinct from conventional, parasthesia-based SCS. A recent study reported that kEFS hyperpolarizes spinal neurons, yet this potentially transformative mode of action contradicts previous evidence that kEFS induces depolarization and was based on patch clamp recordings whose accuracy in the presence of kEFS has not been verified. ObjectivesWe sought to elucidate the basis for kEFS-induced hyperpolarization and to validate the effects of kEFS observed in patch clamp recordings by comparing with independent optical methods. MethodsUsing patch clamp electrophysiology and voltage-sensitive dye (VSD) imaging, we measured the response to kEFS applied in vitro to hippocampal and spinal neurons. ResultsThe kEFS-induced hyperpolarization observed with current clamp recordings was corroborated by VSD imaging and rheobase measurements in patched neurons. However, no hyperpolarization was observed when imaging unpatched neurons or when recording with a voltage-follower amplifier rather than with a patch clamp amplifier (PCA). We found that EFS induced an artifactual current in PCAs that was injected back into current clamped neurons. The artifactual current induced by single, charge-balanced EFS pulses caused modest hyperpolarization, but these unitary hyperpolarizations accumulated when EFS pulses were repeated at kilohertz frequencies. ConclusionOur results rule out hyperpolarization as the mechanism underlying kEFS-mediated analgesia and highlight the risk of recording artifacts caused by extracellular electrical stimulation.

Effects of prasugrel on membrane potential and contractile activity of rat ventricular myocytes.

Though prasugrel is one of the important P2Y12 inhibitors currently in use for antiplatelet therapy, its potential effects on contractility and electrical activity of ventricular myocytes have not yet been investigated. Hence this study was designed to study the impact of prasugrel on contractile function and membrane potential of isolated ventricular myocytes. Freshly isolated rat ventricular myocytes were used in this study. Myocyte contraction was measured during electrical stimulation of cardiomyocytes and the action potential (AP) recordings were obtained with current clamp mode of the patch-clamp amplifier. AP duration and fractional shortening of ventricular myocytes did not show any change with the administration of 1μM of prasugrel. However, remarkable depolarization of resting membrane potential followed by apparent fibrillation episodes was detected in the cardiomyocytes. Similar events were observed in the contractile activity of myocytes during field stimulation. Also, a higher concentration of prasugrel (10μM) elicited repeated fibrillations, which disappeared after washout or nitric oxide synthase (NOS) inhibition with L-NAME. In contrast, the same concentration of ticagrelor, another P2Y12 inhibitor did not induce fibrillation events though it decreased the contractility of ventricular myocytes significantly. The perfusion of ventricular myocytes with L-NAME did not alter the negative inotropic effect of ticagrelor. Prasugrel, a widely used antithrombotic agent, may induce depolarization in the membrane potential of myocytes as well as fibrillation via NO mediated pathway.

Nanopore Logic Operation with DNA to RNA Transcription in a Droplet System.

This paper describes an AND logic operation with amplification and transcription from DNA to RNA, using T7 RNA polymerase. All four operations, (0 0) to (1 1), with an enzyme reaction can be performed simultaneously, using four-droplet devices that are directly connected to a patch-clamp amplifier. The output RNA molecule is detected using a biological nanopore with single-molecule translocation. Channel current recordings can be obtained using the enzyme solution. The integration of DNA logic gates into electrochemical devices is necessary to obtain output information in a human-recognizable form. Our method will be useful for rapid and confined DNA computing applications, including the development of programmable diagnostic devices.

Collisions of Ir Oxide Nanoparticles with Carbon Nanopipettes: Experiments with One Nanoparticle.

Investigating the collisions of individual metal nanoparticles (NPs) with electrodes can provide new insights into their electrocatalytic behavior, mass transport, and interactions with surfaces. Here we report a new experimental setup for studying NP collisions based on the use of carbon nanopipettes to enable monitoring multiple collision events involving the same NP captured inside the pipet cavity. A patch clamp amplifier capable of measuring pA-range currents on the microsecond time scale with a very low noise and stable background was used to record the collision transients. The analysis of current transients produced by oxidation of hydrogen peroxide at one IrOx NP provided information about the origins of deactivation of catalytic NPs and the effects of various experimental conditions on the collision dynamics. High-resolution TEM of carbon pipettes was used to attain better understanding of the NP capture and collisions.