Automated patch clamp

Last updated

Automated patch clamping is beginning [1] to replace manual patch clamping as a method to measure the electrical activity of individual cells. Different techniques are used to automate patch clamp recordings from cells in cell culture and in vivo. This work has been ongoing since the late 1990s by research labs and companies trying to reduce its complexity and cost of patch clamping manually. Patch clamping for a long time was considered an art form and is still very time consuming and tedious, especially in vivo. The automation techniques try to reduce user error and variability in obtaining quality electrophysiology recordings from single cells.

Contents

Manual patch clamp

Animation showing the gigasealing process. The pipette approaches the cell and a plume of liquid flowing out of the pipette makes a small dimple on the surface of the cell. When the resistance has increased enough, a small amount of suction is applied to the pipette which draws the cell membrane into contact with the pipette tip. This creates the gigaohm seal characteristic of a patch clamp recording. Patch Clamp Resistance Animation.gif
Animation showing the gigasealing process. The pipette approaches the cell and a plume of liquid flowing out of the pipette makes a small dimple on the surface of the cell. When the resistance has increased enough, a small amount of suction is applied to the pipette which draws the cell membrane into contact with the pipette tip. This creates the gigaohm seal characteristic of a patch clamp recording.

The traditional manual method to patch clamp using glass pipettes was developed by Erwin Neher and Bert Sakmann and required a highly skilled technician. The technician would position the glass pipette near a cell and apply the appropriate suction to create an electrical seal between the pipette and the cell membrane. This seal ensures a quality recording by preventing any current from leaking out between the tip of the pipette and the cell membrane. This seal is made when the membrane of the cell chemically binds with the tip of the pipette so that the inside of the pipette is only connected to the cytoplasm of the cell. This membrane-glass connection or seal is called a "gigaseal". [2]

The technician traditionally used their mouth to provide the precise pressures required to seal it to the cell. In addition to controlling the pressure, the technician must also position the pipette at precisely the correct distance from the cell so that the membrane will seal with it. Using a micromanipulator, the pipette is moved towards the cell until the technician sees a change in the electrical resistance between the fluid inside of the pipette and the surrounding fluid (see animation). This typically requires 3–12 months of training before a technician is able to reliably record from cells. The technician is essentially performing a balancing act trying to watch and manipulate several systems simultaneously (motion, pressure, and electrical signals). Unless each portion of the process is performed accurately and with the right timing, the seal will not be formed properly and the technician will have to replace the pipette and start over.

These challenges reduce the number of recordings a technician can obtain, and significantly increase the cost. Automation seeks to reduce the time, complexity and cost of manual patch clamping. Improving throughput will also be key to enabling high throughput patch-seq for simultaneously combining electrophysiology, morphology, and transcriptomic properties of neurons at the scale comparable to other sequencing methods [3]

Automation systems

The automation technique varies, depending on the surrounding environment of the cells. For cells in vivo, this typically means that the cells are in the brain and surrounded by other cells. This environment also contains blood vessels, dendrites, axons, and glial cells which make it harder to form a gigaseal by clogging the 1-2μm diameter pipette tip. Here, the precise control of pressure and position at the pipette tip plays a big role in preventing clogging, and detecting whether a cell is near the tip of the pipette, as discussed above.

Cells in vitro can be suspended in a fluid, made to adhere to a culture dish, or remain part of a piece of tissue that has been removed from the animal. These environments typically don't have to compensate for motion of the tissue due to the heartbeat or breathing of an animal. In the case of cells in suspension, the pipette is completely replaced with a microchip with holes that can create gigaseals and measure the electrical activity. Clogging is also less problematic for cells or tissue in culture dishes because the cells and pipette can be seen through a microscope which helps the technician avoid everything but the cell of interest.

Each of these automated systems must perform several tasks. It must position the cell next to the tip of a pipette, or some other device with a 1-2μm hole, control the pressure at the hole, and control the voltage inside the cell.

In vivo

One example of in vivo patch clamping was shown by Kodandaramaiah, et al. [4] In this case the pressure control consisted of a set of electronic valves and electronic pressure regulators to provide three pressures that were previously provided by a technician (high pressure 800-1000mbar, low pressure 20-30mbar, and a small vacuum 15-150mbar). Three electronic valves switched between the three pressures and atmospheric pressure. The high pressure was used to prevent clogging the pipette, the low pressure was used when searching for cells, and the vacuum was used to help the gigasealing process. These were all controlled by a computer, to select among the pressures as the resistance at the tip of the pipette changed.

The manual position control in this case was replaced by a computer controlled piezoelectric micromanipulator that moved the pipette in discrete 2-3μm steps into the tissue until it made contact with a cell. This precision control is much more accurate and repeatable than manual positioning and doesn't require an operator.

The computer also calculates and tracks the change in the electrical resistance as the pipette makes contact with the cell. It sends a voltage signal in the form of a square wave down the pipette which either exits the end of the pipette or is blocked by the cell membrane. When the membrane blocks it, the computer stops the motion of the pipette and applies suction to form the gigaseal. This automation eliminates the decision-making a technician had to perform, and unlike a technician, the computer can perform these tasks tirelessly and with greater precision.

All of these steps are performed in the same logical sequence as manual patch clamping, but don't require extensive training to perform, and are completely controlled by the computer. This reduces the expense required to obtain patch clamp recordings and increases the repeatability and robustness of recording in the living brain.

In suspension

Schematic of a patch clamp system using a droplet suspension culture and gravity to position the cells above the pipette. Suction inside the pipette draws the cells to the tip of the pipette which then forms the gigaseal. Patch Clamp Droplet.svg
Schematic of a patch clamp system using a droplet suspension culture and gravity to position the cells above the pipette. Suction inside the pipette draws the cells to the tip of the pipette which then forms the gigaseal.
A schematic of a patch clamp chip showing a gigaseal, whole cell recording configuration, and the ion channel and whole cell currents. Patch Clamp Chip.svg
A schematic of a patch clamp chip showing a gigaseal, whole cell recording configuration, and the ion channel and whole cell currents.

Many types of systems have been developed for patch clamping cells in suspension cultures. One system uses a traditional pipette and cells in a droplet suspension culture to obtain patch clamp recordings (see figure). This has the added benefit of using traditional pipette fabrication systems that heat a glass capillary and pull it lengthwise to create the tapered tip used in patch clamping.

More common automation systems for suspensions cultures use microchips with tiny (1-2μm) holes in a planar substrate instead of pipettes to create the gigaseal and record from single cells. Patch chips were developed in the early 2000s as a result of the improvement of microfabrication technologies developed by the semiconductor industry. Chips are typically made from silicon, glass, PDMS, polymide. The patch chip systems are usually more complex and expensive but have the added benefit of parallel and hands-free operation. [5] [6] [7]

Ordinarily, neurons don't grow in suspension cultures, but other cell types can. Some can be transfected with genes to create the membrane ion channels of interest. This means that a cell that normally doesn't have electrical activity can grow ion channels in its membrane that will generate ionic currents. Because the cells are dissociated from one another in suspension cultures, the ionic currents in a single cell can be measured with precision. This allows researchers to study ion channel behavior in more controlled environments without currents from other cells interfering, as usually occurs in neural networks. This is particularly useful in drug screening studies where the target is a specific protein. [8] Because handling cells in suspension is much easier than handling cells in culture or in vivo, patch clamp recordings can be obtained much faster and more reliably this way, which increases productivity, making the screening of thousands of compounds possible. [9]

Neurons derived from stem cells cultured adherently can be lifted into suspension and have been successfully used on planar patch clamp devices. [10] Ion channels such as voltage-gated sodium channels, voltage-gated potassium channels and ionotropic ligand-gated ion channels opened by the ligand GABA were recorded from these cells using automated and manual patch clamp. [11]

In culture

There are many in vitro methods for the automated patch clamping of cultured cells or slices of brain tissue.

One uses a patch chip, like those discussed above, along with surface treatments that cause the cultured cells to migrate to the orifices where the gigaseal is formed as they grow. [12] By allowing the neurons to grow in culture, they form networks spontaneously, like those in the brain, which is more like the natural tissues than isolated cells in suspension.

In another method, cells are removed from an animal and cultured on the patch chip for 2–4 hours as they spontaneously form gigaseals with polyimide and PDMS patch chips [13] This system requires no external equipment to form gigaseals.

Another technique automates the positioning of patch clamping cells in cultures. It uses a nanopipette on a precise, piezo-actuated stage to scan a surface within a culture dish. As it scans, it maintains a constant electrical capacitance between the tip of the pipette and the surface or cells beneath it by moving it up and down. (As it moves close to a cell, the capacitance increases, so the actuator moves the pipette away, and vice versa.) This give a precise topographical mapping of the surface within the culture dish. After the cells have been mapped, the computer moves the pipette over to a selected cell and lowers it to form a gigaseal with it. [14]

Another technique simply automates the business of carefully making contact with cells. The operator positions a pipette over the sample and then lets the automated software take over, lowering the pipette and seeking to detect an increase in resistance on the pipette as it makes contact with a cell. At this point the process ends, and a technician creates the gigaseal manually.

Development and acceptance

Patch-clamp automation instrumentation became commercially available in 2003. Due to the initial high cost this near to 20 years old technology was originally intended to serve the biotech and pharmaceutical industries but for the last years its presence has been growing in academia and nonprofit settings, given its increasing proven technical reliability and relative accessibility in cost. A growing number of universities and other academic institutions now have laboratories and core facilities equipped with patch-clamp automated apparatuses in connection and coexisting with other associated or complementary technologies and methods. [15] The acceptance and recognition of automation patch-clamp electrophysiology is reflected in the exponential growth of scientific literature published with results obtained with this revolutionary new technology [16]

Related Research Articles

Neuron Electrically excitable cell that communicates via synapses

A neuron or nerve cell is a electrically excitable cell that communicates with other cells via specialized connections called synapses. It is the main component of nervous tissue in all animals except sponges and placozoa. Plants and fungi do not have nerve cells.

Electrophysiology Study of the electrical properties of biological cells and tissues.

Electrophysiology is the branch of physiology that studies the electrical properties of biological cells and tissues. It involves measurements of voltage changes or electric current or manipulations on a wide variety of scales from single ion channel proteins to whole organs like the heart. In neuroscience, it includes measurements of the electrical activity of neurons, and, in particular, action potential activity. Recordings of large-scale electric signals from the nervous system, such as electroencephalography, may also be referred to as electrophysiological recordings. They are useful for electrodiagnosis and monitoring.

Action potential Process by which neurons communicate with each other by changes in their membrane potentials

In physiology, an action potential (AP) occurs when the membrane potential of a specific cell location rapidly rises and falls: this depolarization then causes adjacent locations to similarly depolarize. Action potentials occur in several types of animal cells, called excitable cells, which include neurons, muscle cells, endocrine cells and in some plant cells.

A spheroplast is a microbial cell from which the cell wall has been almost completely removed, as by the action of penicillin or lysozyme. According to some definitions, the term is used to describe Gram-negative bacteria. According to other definitions, the term also encompasses yeasts. The name spheroplast stems from the fact that after the microbe's cell wall is digested, membrane tension causes the cell to acquire a characteristic spherical shape. Spheroplasts are osmotically fragile, and will lyse if transferred to a hypotonic solution.

Hyperpolarization is a change in a cell's membrane potential that makes it more negative. It is the opposite of a depolarization. It inhibits action potentials by increasing the stimulus required to move the membrane potential to the action potential threshold.

Stimulus (physiology) Detectable change in the internal or external surroundings

In physiology, a stimulus is a detectable change in the physical or chemical structure of an organism's internal or external environment. The ability of an organism or organ to detect external stimuli, so that an appropriate reaction can be made, is called sensitivity (excitability). Sensory receptors can receive information from outside the body, as in touch receptors found in the skin or light receptors in the eye, as well as from inside the body, as in chemoreceptors and mechanoreceptors. When a stimulus is detected by a sensory receptor, it can elicit a reflex via stimulus transduction. An internal stimulus is often the first component of a homeostatic control system. External stimuli are capable of producing systemic responses throughout the body, as in the fight-or-flight response. In order for a stimulus to be detected with high probability, its level of strength must exceed the absolute threshold; if a signal does reach threshold, the information is transmitted to the central nervous system (CNS), where it is integrated and a decision on how to react is made. Although stimuli commonly cause the body to respond, it is the CNS that finally determines whether a signal causes a reaction or not.

Threshold potential

In electrophysiology, the threshold potential is the critical level to which a membrane potential must be depolarized to initiate an action potential. In neuroscience, threshold potentials are necessary to regulate and propagate signaling in both the central nervous system (CNS) and the peripheral nervous system (PNS).

Voltage clamp

The voltage clamp is an experimental method used by electrophysiologists to measure the ion currents through the membranes of excitable cells, such as neurons, while holding the membrane voltage at a set level. A basic voltage clamp will iteratively measure the membrane potential, and then change the membrane potential (voltage) to a desired value by adding the necessary current. This "clamps" the cell membrane at a desired constant voltage, allowing the voltage clamp to record what currents are delivered. Because the currents applied to the cell must be equal to the current going across the cell membrane at the set voltage, the recorded currents indicate how the cell reacts to changes in membrane potential. Cell membranes of excitable cells contain many different kinds of ion channels, some of which are voltage-gated. The voltage clamp allows the membrane voltage to be manipulated independently of the ionic currents, allowing the current–voltage relationships of membrane channels to be studied.

Patch clamp

The patch clamp technique is a laboratory technique in electrophysiology used to study ionic currents in individual isolated living cells, tissue sections, or patches of cell membrane. The technique is especially useful in the study of excitable cells such as neurons, cardiomyocytes, muscle fibers, and pancreatic beta cells, and can also be applied to the study of bacterial ion channels in specially prepared giant spheroplasts.

The sucrose gap technique is used to create a conduction block in nerve or muscle fibers. A high concentration of sucrose is applied to the extracellular space, which prevents the correct opening and closing of sodium and potassium channels, increasing resistance between two groups of cells. It was originally developed by Robert Stämpfli for recording action potentials in nerve fibers, and is particularly useful for measuring irreversible or highly variable pharmacological modifications of channel properties since untreated regions of membrane can be pulled into the node between the sucrose regions.

In neuroscience, single-unit recordings provide a method of measuring the electro-physiological responses of a single neuron using a microelectrode system. When a neuron generates an action potential, the signal propagates down the neuron as a current which flows in and out of the cell through excitable membrane regions in the soma and axon. A microelectrode is inserted into the brain, where it can record the rate of change in voltage with respect to time. These microelectrodes must be fine-tipped, low-impedance conductors; they are primarily glass micro-pipettes, metal microelectrodes made of platinum, tungsten, iridium or even iridium oxide. Microelectrodes can be carefully placed close to the cell membrane, allowing the ability to record extracellularly.

Scanning ion-conductance microscopy

Scanning ion-conductance microscopy (SICM) is a scanning probe microscopy technique that uses an electrode as the probe tip. SICM allows for the determination of the surface topography of micrometer and even nanometer-range structures in aqueous media conducting electrolytes. The samples can be hard or soft, are generally non-conducting, and the non-destructive nature of the measurement allows for the observation of living tissues and cells, and biological samples in general.

A microelectrode is an electrode used in electrophysiology either for recording neural signals or for the electrical stimulation of nervous tissue. Pulled glass pipettes with tip diameters of 0.5 μm or less are usually filled with 3 molars potassium chloride solution as the electrical conductor. When the tip penetrates a cell membrane the lipids in the membrane seal onto the glass, providing an excellent electrical connection between the tip and the interior of the cell, which is apparent because the microelectrode becomes electrically negative compared to the extracellular solution. There are also microelectrodes made with insulated metal wires, made from inert metals with high Young modulus such as tungsten, stainless steel, or platinum-iridium alloy and coated with glass or polymer insulator with exposed conductive tips. These are mostly used for recording from the external side of the cell membrane. More recent advances in lithography have produced silicon-based microelectrodes.

Subthreshold membrane potential oscillations

Subthreshold membrane potential oscillations are membrane oscillations that do not directly trigger an action potential since they do not reach the necessary threshold for firing. However, they may facilitate sensory signal processing.

Microelectrode arrays (MEAs) are devices that contain multiple microelectrodes through which neural signals are obtained or delivered, essentially serving as neural interfaces that connect neurons to electronic circuitry. There are two general classes of MEAs: implantable MEAs, used in vivo, and non-implantable MEAs, used in vitro.

Dendritic spike

In neurophysiology, a dendritic spike refers to an action potential generated in the dendrite of a neuron. Dendrites are branched extensions of a neuron. They receive electrical signals emitted from projecting neurons and transfer these signals to the cell body, or soma. Dendritic signaling has traditionally been viewed as a passive mode of electrical signaling. Unlike its axon counterpart which can generate signals through action potentials, dendrites were believed to only have the ability to propagate electrical signals by physical means: changes in conductance, length, cross sectional area, etc. However, the existence of dendritic spikes was proposed and demonstrated by W. Alden Spencer, Eric Kandel, Rodolfo Llinás and coworkers in the 1960s and a large body of evidence now makes it clear that dendrites are active neuronal structures. Dendrites contain voltage-gated ion channels giving them the ability to generate action potentials. Dendritic spikes have been recorded in numerous types of neurons in the brain and are thought to have great implications in neuronal communication, memory, and learning. They are one of the major factors in long-term potentiation.

Mechanosensitive channels, mechanosensitive ion channels or stretch-gated ion channels (not to be confused with mechanoreceptors). They are present in the membranes of organisms from the three domains of life: bacteria, archaea, and eukarya. They are the sensors for a number of systems including the senses of touch, hearing and balance, as well as participating in cardiovascular regulation and osmotic homeostasis (e.g. thirst). The channels vary in selectivity for the permeating ions from nonselective between anions and cations in bacteria, to cation selective allowing passage Ca2+, K+ and Na+ in eukaryotes, and highly selective K+ channels in bacteria and eukaryotes.

The channelome, sometimes called the "ion channelome", is the complete set of ion channels and porins expressed in a biological tissue or organism. It is analogous to the genome, the metabolome, the proteome, and the microbiome. Characterization of the ion channelome, referred to as channelomics, is a branch of physiology, biophysics, neuroscience, and pharmacology, with particular attention paid to gene expression. It can be performed by a variety of techniques, including patch clamp electrophysiology, PCR, and immunohistochemistry. Channelomics is being used to screen and discover new medicines.

Single-Entity Electrochemistry (SEE) refers to the electroanalysis of an individual unit of interest. A unique feature of SEE is that it unifies multiple different branches of electrochemistry. Single-Entity Electrochemistry pushes the bounds of the field as it can measure entities on a scale of 100 microns to angstroms. Single-Entity Electrochemistry is important because it gives the ability to view how a single molecule, or cell, or "thing" affects the bulk response, and thus the chemistry that might have gone unknown otherwise. The ability to monitor the movement of one electron or ion from one unit to another is valuable, as many vital reactions and mechanisms undergo this process. Electrochemistry is well suited for this measurement due to its incredible sensitivity. Single-Entity Electrochemistry can be used to investigate nanoparticles, wires, vesicles, nanobubbles, nanotubes, cells, and viruses, and other small molecules and ions. Single-entity electrochemistry has been successfully used to determine the size distribution of particles as well as the number of particles present inside a vesicle or other similar structures

Patch-sequencing

Patch-sequencing (patch-seq) is a method designed for tackling specific problems involved in characterizing neurons. As neural tissues are one of the most transcriptomically diverse populations of cells, classifying neurons into cell types in order to understand the circuits they form is a major challenge for neuroscientists. Combining classical classification methods with single cell RNA-sequencing post-hoc has proved to be difficult and slow. By combining multiple data modalities such as electrophysiology, sequencing and microscopy, Patch-seq allows for neurons to be characterized in multiple ways simultaneously. It currently suffers from low throughput relative to other sequencing methods mainly due to the manual labor involved in achieving a successful patch-clamp recording on a neuron. Investigations are currently underway to automate patch-clamp technology which will improve the throughput of patch-seq as well.

References

  1. ↑ Wood C, Williams C, Waldron GJ (May 2004). "Patch clamping by numbers". Drug Discovery Today. 9 (10): 434–41. doi:10.1016/S1359-6446(04)03064-8. PMID   15109948.
  2. ↑ Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (August 1981). "Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches". Pflügers Archiv. 391 (2): 85–100. CiteSeerX   10.1.1.456.107 . doi:10.1007/BF00656997. PMID   6270629. S2CID   12014433.
  3. ↑ Tripathy, Shreejoy J.; Toker, Lilah; Bomkamp, Claire; Mancarci, B. Ogan; Belmadani, Manuel; Pavlidis, Paul (2018-10-08). "Assessing Transcriptome Quality in Patch-Seq Datasets". Frontiers in Molecular Neuroscience. 11: 363. doi: 10.3389/fnmol.2018.00363 . ISSN   1662-5099. PMC   6187980 . PMID   30349457.
  4. ↑ Kodandaramaiah SB, Franzesi GT, Chow BY, Boyden ES, Forest CR (June 2012). "Automated whole-cell patch-clamp electrophysiology of neurons in vivo". Nature Methods. 9 (6): 585–7. doi:10.1038/nmeth.1993. PMC   3427788 . PMID   22561988.
  5. ↑ Jones KA, Garbati N, Zhang HM, Large CH (2009). "Automated Patch Clamping Using the QPatch". High Throughput Screening. Methods in Molecular Biology. 565. pp. 209–23. doi:10.1007/978-1-60327-258-2_10. ISBN   978-1-60327-257-5. PMID   19551364.
  6. ↑ Chen CY, Tu TY, Chen CH, Jong DS, Wo AM (August 2009). "Patch clamping on plane glass-fabrication of hourglass aperture and high-yield ion channel recording". Lab on a Chip. 9 (16): 2370–80. doi:10.1039/B901025D. PMID   19636469.
  7. ↑ Obergrussberger A, Stölzle-Feix S, Becker N, Brüggemann A, Fertig N, Möller C (2015). "Novel screening techniques for ion channel targeting drugs". Channels. 9 (6): 367–75. doi:10.1080/19336950.2015.1079675. PMC   4850050 . PMID   26556400.
  8. ↑ Dunlop J, Bowlby M, Peri R, Vasilyev D, Arias R (April 2008). "High-throughput electrophysiology: an emerging paradigm for ion-channel screening and physiology". Nature Reviews. Drug Discovery. 7 (4): 358–68. doi:10.1038/nrd2552. PMID   18356919. S2CID   9294213.
  9. ↑ Obergrussberger A, Brüggemann A, Goetze TA, Rapedius M, Haarmann C, Rinke I, et al. (December 2016). "Automated Patch Clamp Meets High-Throughput Screening: 384 Cells Recorded in Parallel on a Planar Patch Clamp Module". Journal of Laboratory Automation. 21 (6): 779–793. doi: 10.1177/2211068215623209 . PMID   26702021.
  10. ↑ Haythornthwaite A, Stoelzle S, Hasler A, Kiss A, Mosbacher J, George M, et al. (October 2012). "Characterizing human ion channels in induced pluripotent stem cell-derived neurons". Journal of Biomolecular Screening. 17 (9): 1264–72. doi: 10.1177/1087057112457821 . PMID   22923790.
  11. ↑ Haythornthwaite A, Stoelzle S, Hasler A, Kiss A, Mosbacher J, George M, et al. (October 2012). "Characterizing human ion channels in induced pluripotent stem cell-derived neurons". Journal of Biomolecular Screening. 17 (9): 1264–72. doi: 10.1177/1087057112457821 . PMID   22923790.
  12. ↑ Py C, Martina M, Diaz-Quijada GA, Luk CC, Martinez D, Denhoff MW, et al. (2011). "From understanding cellular function to novel drug discovery: the role of planar patch-clamp array chip technology". Frontiers in Pharmacology. 2: 51. doi: 10.3389/fphar.2011.00051 . PMC   3184600 . PMID   22007170.
  13. ↑ Martinez D, Py C, Denhoff MW, Martina M, Monette R, Comas T, et al. (December 2010). "High-fidelity patch-clamp recordings from neurons cultured on a polymer microchip". Biomedical Microdevices. 12 (6): 977–85. CiteSeerX   10.1.1.362.9850 . doi:10.1007/s10544-010-9452-z. PMID   20694518. S2CID   14441945.
  14. ↑ Gorelik J, Gu Y, Spohr HA, Shevchuk AI, Lab MJ, Harding SE, et al. (December 2002). "Ion channels in small cells and subcellular structures can be studied with a smart patch-clamp system". Biophysical Journal. 83 (6): 3296–303. Bibcode:2002BpJ....83.3296G. doi:10.1016/S0006-3495(02)75330-7. PMC   1302405 . PMID   12496097.
  15. ↑ Picones A, Loza-Huerta A, Segura-Chama P, Lara-Figueroa CO (2016). "Contribution of Automated Technologies to Ion Channel Drug Discovery". Advances in Protein Chemistry and Structural Biology. 104: 357–378. doi:10.1016/bs.apcsb.2016.01.002. ISBN   9780128048269. PMID   27038379.
  16. ↑ Picones A (2015). "Ion channels as medicinal targets of biological toxins: the impact of automated patch-clamp electrophysiology". Current Topics in Medicinal Chemistry. 15 (7): 631–7. doi:10.2174/1568026615666150309145928. PMID   25751269.
This page is based on this Wikipedia article
Text is available under the CC BY-SA 4.0 license; additional terms may apply.
Images, videos and audio are available under their respective licenses.