![Single channel](https://upload.wikimedia.org/wikipedia/commons/8/8d/Single_channel.png)
The patch clamp technique is a
laboratory technique
A laboratory (; ; colloquially lab) is a facility that provides controlled conditions in which scientific or technological research, experiments, and measurement may be performed. Laboratory services are provided in a variety of settings: physici ...
in
electrophysiology
Electrophysiology (from Greek , ''ēlektron'', "amber" etymology of "electron"">Electron#Etymology">etymology of "electron" , ''physis'', "nature, origin"; and , '' -logia'') is the branch of physiology that studies the electrical properties of b ...
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
neuron
A neuron, neurone, or nerve cell is an electrically excitable cell that communicates with other cells via specialized connections called synapses. The neuron is the main component of nervous tissue in all animals except sponges and placozoa. N ...
s,
cardiomyocyte
Cardiac muscle (also called heart muscle, myocardium, cardiomyocytes and cardiac myocytes) is one of three types of vertebrate muscle tissues, with the other two being skeletal muscle and smooth muscle. It is an involuntary, striated muscle that ...
s,
muscle fiber
A muscle cell is also known as a myocyte when referring to either a cardiac muscle cell (cardiomyocyte), or a smooth muscle cell as these are both small cells. A skeletal muscle cell is long and threadlike with many nuclei and is called a muscl ...
s, and
pancreatic
The pancreas is an Organ (anatomy), organ of the digestive system and endocrine system of vertebrates. In humans, it is located in the abdominal cavity, abdomen behind the stomach and functions as a gland. The pancreas is a mixed or heterocrine ...
beta cell
Beta cells (β-cells) are a type of cell found in pancreatic islets that synthesize and secrete insulin and amylin. Beta cells make up 50–70% of the cells in human islets. In patients with Type 1 diabetes, beta-cell mass and function are dimini ...
s, and can also be applied to the study of
bacteria
Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one biological cell. They constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria were among ...
l ion channels in specially prepared giant
spheroplasts
A spheroplast (or sphaeroplast in British usage) 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 ...
.
Patch clamping can be performed using the
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 iterativ ...
technique. In this case, the voltage across the cell membrane is controlled by the experimenter and the resulting currents are recorded. Alternatively, the
current clamp
In electrical and electronic engineering, a current clamp, also known as current probe, is an electrical device with jaws which open to allow clamping around an electrical conductor. This allows measurement of the current in a conductor without t ...
technique can be used. In this case, the current passing across the membrane is controlled by the experimenter and the resulting changes in voltage are recorded, generally in the form of
action potential
An action potential 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, ...
s.
Erwin Neher
Erwin Neher (; ; born 20 March 1944) is a German biophysicist, specializing in the field of cell physiology. For significant contribution in the field, in 1991 he was awarded, along with Bert Sakmann, the Nobel Prize in Physiology or Medicine for ...
and
Bert Sakmann
Bert Sakmann (; born 12 June 1942) is a German cell physiologist. He shared the Nobel Prize in Physiology or Medicine with Erwin Neher in 1991 for their work on "the function of single ion channels in cells," and the invention of the patch cla ...
developed the patch clamp in the late 1970s and early 1980s. This discovery made it possible to record the currents of single ion channel molecules for the first time, which improved understanding of the involvement of channels in fundamental cell processes such as action potentials and nerve activity. Neher and Sakmann received the
Nobel Prize in Physiology or Medicine
The Nobel Prize in Physiology or Medicine is awarded yearly by the Nobel Assembly at the Karolinska Institute for outstanding discoveries in physiology or medicine. The Nobel Prize is not a single prize, but five separate prizes that, accord ...
in 1991 for this work.
Basic technique
Set-up
![Patch Clamp Rig classic](https://upload.wikimedia.org/wikipedia/commons/3/35/Patch_Clamp_Rig_classic.jpg)
During a patch clamp recording, a hollow glass tube known as a
micropipette
A pipette (sometimes spelled as pipett) is a laboratory tool commonly used in chemistry, biology and medicine to transport a measured volume of liquid, often as a media dispenser. Pipettes come in several designs for various purposes with diffe ...
or patch pipette filled with an electrolyte solution and a recording
electrode
An electrode is an electrical conductor used to make contact with a nonmetallic part of a circuit (e.g. a semiconductor, an electrolyte, a vacuum or air). Electrodes are essential parts of batteries that can consist of a variety of materials de ...
connected to an amplifier is brought into contact with the membrane of an
isolated cell. Another electrode is placed in a bath surrounding the cell or tissue as a reference
ground
Ground may refer to:
Geology
* Land, the surface of the Earth not covered by water
* Soil, a mixture of clay, sand and organic matter present on the surface of the Earth
Electricity
* Ground (electricity), the reference point in an electrical c ...
electrode. An electrical circuit can be formed between the recording and reference electrode with the cell of interest in between.
![Patch clamp schematic 1](https://upload.wikimedia.org/wikipedia/commons/6/60/Patch_clamp_schematic_1.png)
The solution filling the patch pipette might match the ionic composition of the bath solution, as in the case of cell-attached recording, or match the
cytoplasm
In cell biology, the cytoplasm is all of the material within a eukaryotic cell, enclosed by the cell membrane, except for the cell nucleus. The material inside the nucleus and contained within the nuclear membrane is termed the nucleoplasm. The ...
, for whole-cell recording. The solution in the bath solution may match the physiological extracellular solution, the cytoplasm, or be entirely non-physiological, depending on the experiment to be performed. The researcher can also change the content of the bath solution (or less commonly the pipette solution) by adding ions or drugs to study the ion channels under different conditions.
Depending on what the researcher is trying to measure, the diameter of the pipette tip used may vary, but it is usually in the
micrometer Micrometer can mean:
* Micrometer (device), used for accurate measurements by means of a calibrated screw
* American spelling of micrometre
The micrometre ( international spelling as used by the International Bureau of Weights and Measures; ...
range.
This small size is used to enclose a
cell membrane
The cell membrane (also known as the plasma membrane (PM) or cytoplasmic membrane, and historically referred to as the plasmalemma) is a biological membrane that separates and protects the interior of all cells from the outside environment ( ...
surface area or "patch" that often contains just one or a few ion channel molecules.
This type of electrode is distinct from the "sharp microelectrode" used to puncture cells in traditional
intracellular recordings, in that it is sealed onto the surface of the cell membrane, rather than inserted through it.
![Patch clamp schematic 2](https://upload.wikimedia.org/wikipedia/commons/4/4e/Patch_clamp_schematic_2.png)
In some experiments, the micropipette tip is heated in a microforge to produce a smooth surface that assists in forming a high
resistance seal with the cell membrane. To obtain this high resistance seal, the micropipette is pressed against a cell membrane and suction is applied. A portion of the cell membrane is suctioned into the pipette, creating an
omega
Omega (; capital: Ω, lowercase: ω; Ancient Greek ὦ, later ὦ μέγα, Modern Greek ωμέγα) is the twenty-fourth and final letter in the Greek alphabet. In the Greek numeric system/isopsephy (gematria), it has a value of 800. The wo ...
-shaped area of membrane which, if formed properly, creates a resistance in the 10–100
gigaohms range, called a "gigaohm seal" or "gigaseal".
The high resistance of this seal makes it possible to isolate electronically the currents measured across the membrane patch with little competing
noise
Noise is unwanted sound considered unpleasant, loud or disruptive to hearing. From a physics standpoint, there is no distinction between noise and desired sound, as both are vibrations through a medium, such as air or water. The difference arise ...
, as well as providing some mechanical stability to the recording.
Recording
![WholeCellPatchClamp-03](https://upload.wikimedia.org/wikipedia/commons/4/41/WholeCellPatchClamp-03.jpg)
Many patch clamp amplifiers do not use true
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 iterativ ...
circuitry, but instead are
differential amplifier
A differential amplifier is a type of electronic amplifier that amplifies the difference between two input voltages but suppresses any voltage common to the two inputs. It is an analog circuit with two inputs V_\text^- and V_\text^+ and one outpu ...
s that use the bath electrode to set the zero current (ground) level. This allows a researcher to keep the voltage constant while observing changes in
current
Currents, Current or The Current may refer to:
Science and technology
* Current (fluid), the flow of a liquid or a gas
** Air current, a flow of air
** Ocean current, a current in the ocean
*** Rip current, a kind of water current
** Current (stre ...
. To make these recordings, the patch pipette is compared to the ground electrode. Current is then injected into the system to maintain a constant, set voltage. The current that is needed to clamp the voltage is opposite in sign and equal in magnitude to the current through the membrane.
Alternatively, the cell can be
current clamped in whole-cell mode, keeping current constant while observing changes in membrane
voltage
Voltage, also known as electric pressure, electric tension, or (electric) potential difference, is the difference in electric potential between two points. In a static electric field, it corresponds to the work needed per unit of charge to m ...
.
Variations
![Patchmodes](https://upload.wikimedia.org/wikipedia/commons/1/1f/Patchmodes.svg)
Several variations of the basic technique can be applied, depending on what the researcher wants to study. The inside-out and outside-out techniques are called "excised patch" techniques, because the patch is excised (removed) from the main body of the cell. Cell-attached and both excised patch techniques are used to study the behavior of individual ion channels in the section of membrane attached to the electrode.
Whole-cell patch and perforated patch allow the researcher to study the electrical behavior of the entire cell, instead of single channel currents. The whole-cell patch, which enables low-resistance electrical access to the inside of a cell, has now largely replaced
high-resistance microelectrode recording techniques to record currents across the entire cell membrane.
Cell-attached patch
![Cell-Attached Patch Clamp](https://upload.wikimedia.org/wikipedia/commons/7/7a/Cell-Attached_Patch_Clamp.png)
For this method, the pipette is sealed onto the cell membrane to obtain a gigaseal (a seal with electrical resistance on the order of a gigaohm), while ensuring that the cell membrane remains intact. This allows the recording of currents through single, or a few, ion channels contained in the patch of membrane captured by the pipette. By only attaching to the exterior of the cell membrane, there is very little disturbance of the cell structure.
Also, by not disrupting the interior of the cell, any intracellular mechanisms normally influencing the channel will still be able to function as they would physiologically.
Using this method it is also relatively easy to obtain the right configuration, and once obtained it is fairly stable.
For
ligand-gated ion channels
Ligand-gated ion channels (LICs, LGIC), also commonly referred to as ionotropic receptors, are a group of transmembrane ion-channel proteins which open to allow ions such as Na+, K+, Ca2+, and/or Cl− to pass through the membrane in res ...
or channels that are modulated by
metabotropic receptors
A metabotropic receptor, also referred to by the broader term G-protein-coupled receptor, is a type of membrane receptor that initiates a number of metabolic steps to modulate cell activity. The nervous system utilizes two types of receptors: met ...
, the
neurotransmitter
A neurotransmitter is a signaling molecule secreted by a neuron to affect another cell across a synapse. The cell receiving the signal, any main body part or target cell, may be another neuron, but could also be a gland or muscle cell.
Neuro ...
or drug being studied is usually included in the pipette solution, where it can interact with what used to be the external surface of the membrane. The resulting channel activity can be attributed to the drug being used, although it is usually not possible to then change the drug concentration inside the pipette. The technique is thus limited to one point in a
dose response curve
Dose or Dosage may refer to:
Music
* Dose (Gov't Mule album), ''Dose'' (Gov't Mule album), 1998
* Dose (Latin Playboys album), ''Dose'' (Latin Playboys album)
* Dosage (album), ''Dosage'' (album), by the band Collective Soul
* Dose (song), "Dose" ...
per patch. Therefore, the dose response is accomplished using several cells and patches. However,
voltage-gated ion channel
Voltage-gated ion channels are a class of transmembrane proteins that form ion channels that are activated by changes in the electrical membrane potential near the channel. The membrane potential alters the conformation of the channel proteins, ...
s can be clamped successively at different membrane potentials in a single patch. This results in channel activation as a function of voltage, and a complete
I-V (current-voltage) curve can be established in only one patch. Another potential drawback of this technique is that, just as the intracellular pathways of the cell are not disturbed, they cannot be directly modified either.
Inside-out patch
![Inside-Out Patch Clamp](https://upload.wikimedia.org/wikipedia/commons/b/b7/Inside-Out_Patch_Clamp.png)
In the inside-out method, a patch of the membrane is attached to the patch pipette, detached from the rest of the cell, and the
cytosolic surface of the membrane is exposed to the external media, or bath.
One advantage of this method is that the experimenter has access to the intracellular surface of the membrane via the bath and can change the chemical composition of what the inside surface of the membrane is exposed to. This is useful when an experimenter wishes to manipulate the environment at the intracellular surface of single ion channels. For example, channels that are activated by intracellular ligands can then be studied through a range of ligand concentrations.
To achieve the inside-out configuration, the pipette is attached to the cell membrane as in the cell-attached mode, forming a gigaseal, and is then retracted to break off a patch of membrane from the rest of the cell. Pulling off a membrane patch often results initially in the formation of a
vesicle
Vesicle may refer to:
; In cellular biology or chemistry
* Vesicle (biology and chemistry), a supramolecular assembly of lipid molecules, like a cell membrane
* Synaptic vesicle
; In human embryology
* Vesicle (embryology), bulge-like features o ...
of membrane in the pipette tip, because the ends of the patch membrane fuse together quickly after excision. The outer face of the vesicle must then be broken open to enter into inside-out mode; this may be done by briefly taking the membrane through the bath solution/air interface, by exposure to a low
Ca2+ solution, or by momentarily making contact with a droplet of
paraffin Paraffin may refer to:
Substances
* Paraffin wax, a white or colorless soft solid that is used as a lubricant and for other applications
* Liquid paraffin (drug), a very highly refined mineral oil used in cosmetics and for medical purposes
* Alkane ...
or a piece of cured
silicone
A silicone or polysiloxane is a polymer made up of siloxane (−R2Si−O−SiR2−, where R = organic group). They are typically colorless oils or rubber-like substances. Silicones are used in sealants, adhesives, lubricants, medicine, cooking ...
polymer.
Whole-cell recording or whole-cell patch
![Whole Cell Patch Clamp Steps](https://upload.wikimedia.org/wikipedia/commons/9/9c/Whole_Cell_Patch_Clamp_Steps.png)
Whole-cell recordings involve recording currents through multiple channels simultaneously, over a large region of the cell membrane. The electrode is left in place on the cell, as in cell-attached recordings, but more suction is applied to rupture the membrane patch, thus providing access from the interior of the pipette to the
intracellular space
Intracellular space is the interior space of the plasma membrane. It contains about two-thirds of TBW. Cellular rupture may occur if the intracellular space becomes dehydrated, or if the opposite happens, where it becomes too bloated. Thus it ...
of the cell. This provides a means to administer and study how treatments (e.g. drugs) can affect cells in real time.
Once the pipette is attached to the cell membrane, there are two methods of breaking the patch. The first is by applying more suction. The amount and duration of this suction depends on the type of cell and size of the pipette. The other method requires a large current pulse to be sent through the pipette. How much current is applied and the duration of the pulse also depend on the type of cell.
For some types of cells, it is convenient to apply both methods simultaneously to break the patch.
The advantage of whole-cell patch clamp recording over
sharp electrode technique recording is that the larger opening at the tip of the patch clamp electrode provides lower resistance and thus better electrical access to the inside of the cell.
A disadvantage of this technique is that because the volume of the electrode is larger than the volume of the cell, the soluble contents of the cell's interior will slowly be replaced by the contents of the electrode. This is referred to as the electrode
"dialyzing" the cell's contents.
After a while, any properties of the cell that depend on soluble intracellular contents will be altered. The pipette solution used usually approximates the high-
potassium
Potassium is the chemical element with the symbol K (from Neo-Latin ''kalium'') and atomic number19. Potassium is a silvery-white metal that is soft enough to be cut with a knife with little force. Potassium metal reacts rapidly with atmosphe ...
environment of the interior of the cell to minimize any changes this may cause. There is often a period at the beginning of a whole-cell recording when one can take measurements before the cell has been dialyzed.
Outside-out patch
![Outside-Out Patch Clamp](https://upload.wikimedia.org/wikipedia/commons/6/64/Outside-Out_Patch_Clamp.png)
The name "outside-out" emphasizes both this technique's complementarity to the inside-out technique, and the fact that it places the external rather than intracellular surface of the cell membrane on the outside of the patch of membrane, in relation to the patch electrode.
The formation of an outside-out patch begins with a whole-cell recording configuration. After the whole-cell configuration is formed, the electrode is slowly withdrawn from the cell, allowing a bulb of membrane to
bleb out from the cell. When the electrode is pulled far enough away, this bleb will detach from the cell and reform as a convex membrane on the end of the electrode (like a ball open at the electrode tip), with the original outside of the membrane facing outward from the electrode.
As the image at the right shows, this means that the fluid inside the pipette will be simulating the intracellular fluid, while a researcher is free to move the pipette and the bleb with its channels to another bath of solution. While multiple channels can exist in a bleb of membrane, single channel recordings are also possible in this conformation if the bleb of detached membrane is small and only contains one channel.
Outside-out patching gives the experimenter the opportunity to examine the properties of an ion channel when it is isolated from the cell and exposed successively to different solutions on the
extracellular
This glossary of biology terms is a list of definitions of fundamental terms and concepts used in biology, the study of life and of living organisms. It is intended as introductory material for novices; for more specific and technical definitions ...
surface of the membrane. The experimenter can perfuse the same patch with a variety of solutions in a relatively short amount of time, and if the channel is activated by a
neurotransmitter
A neurotransmitter is a signaling molecule secreted by a neuron to affect another cell across a synapse. The cell receiving the signal, any main body part or target cell, may be another neuron, but could also be a gland or muscle cell.
Neuro ...
or drug from the extracellular face, a
dose-response curve can then be obtained.
This ability to measure current through exactly the same piece of membrane in different solutions is the distinct advantage of the outside-out patch relative to the cell-attached method. On the other hand, it is more difficult to accomplish. The longer formation process involves more steps that could fail and results in a lower frequency of usable patches.
Perforated patch
![Perforated Patch Clamp](https://upload.wikimedia.org/wikipedia/commons/b/b3/Perforated_Patch_Clamp.png)
This variation of the patch clamp method is very similar to the whole-cell configuration. The main difference lies in the fact that when the experimenter forms the gigaohm seal, suction is not used to rupture the patch membrane. Instead, the electrode solution contains small amounts of an antifungal or
antibiotic
An antibiotic is a type of antimicrobial substance active against bacteria. It is the most important type of antibacterial agent for fighting bacterial infections, and antibiotic medications are widely used in the treatment and prevention of ...
agent, such as
amphothericin-B,
nystatin
Nystatin, sold under the brandname Mycostatin among others, is an antifungal medication. It is used to treat '' Candida'' infections of the skin including diaper rash, thrush, esophageal candidiasis, and vaginal yeast infections. It may also be ...
, or
gramicidin
Gramicidin, also called gramicidin D, is a mix of ionophoric antibiotics, gramicidin A, B and C, which make up about 80%, 5%, and 15% of the mix, respectively. Each has 2 isoforms, so the mix has 6 different types of gramicidin molecules. They c ...
, which diffuses into the membrane patch and forms small pores in the membrane, providing electrical access to the cell interior.
When comparing the whole-cell and perforated patch methods, one can think of the whole-cell patch as an open door, in which there is complete exchange between molecules in the pipette solution and the cytoplasm. The perforated patch can be likened to a screen door that only allows the exchange of certain molecules from the pipette solution to the cytoplasm of the cell.
Advantages of the perforated patch method, relative to whole-cell recordings, include the properties of the antibiotic pores, that allow equilibration only of small monovalent ions between the patch pipette and the cytosol, but not of larger molecules that cannot permeate through the pores. This property maintains endogenous levels of
divalent ions such as Ca
2+ and signaling molecules such as
cAMP
Camp may refer to:
Outdoor accommodation and recreation
* Campsite or campground, a recreational outdoor sleeping and eating site
* a temporary settlement for nomads
* Camp, a term used in New England, Northern Ontario and New Brunswick to descri ...
. Consequently, one can have recordings of the entire cell, as in whole-cell patch clamping, while retaining most intracellular signaling mechanisms, as in cell-attached recordings. As a result, there is reduced current rundown, and stable perforated patch recordings can last longer than one hour.
Disadvantages include a higher access resistance, relative to whole-cell, due to the partial membrane occupying the tip of the electrode. This may decrease current resolution and increase recording noise. It can also take a significant amount of time for the antibiotic to perforate the membrane (about 15 minutes for amphothericin-B, and even longer for gramicidin and nystatin). The membrane under the electrode tip is weakened by the perforations formed by the antibiotic and can rupture. If the patch ruptures, the recording is then in whole-cell mode, with antibiotic contaminating the inside of the cell.
Loose patch
![Loose Patch Clamp](https://upload.wikimedia.org/wikipedia/commons/0/04/Loose_Patch_Clamp.png)
A loose patch clamp is different from the other techniques discussed here in that it employs a loose seal (low electrical resistance) rather than the tight gigaseal used in the conventional technique. This technique was used as early as the year 1961, as described in a paper by Strickholm on the impedance of a muscle cell's surface,
but received little attention until being brought up again and given a name by Almers, Stanfield, and Stühmer in 1982,
after patch clamp had been established as a major tool of electrophysiology.
To achieve a loose patch clamp on a cell membrane, the pipette is moved slowly towards the cell, until the electrical resistance of the contact between the cell and the pipette increases to a few times greater resistance than that of the electrode alone. The closer the pipette gets to the membrane, the greater the resistance of the pipette tip becomes, but if too close a seal is formed, and it could become difficult to remove the pipette without damaging the cell. For the loose patch technique, the pipette does not get close enough to the membrane to form a gigaseal or a permanent connection, nor to pierce the cell membrane.
The cell membrane stays intact, and the lack of a tight seal creates a small gap through which ions can pass outside the cell without entering the pipette.
A significant advantage of the loose seal is that the pipette that is used can be repeatedly removed from the membrane after recording, and the membrane will remain intact. This allows repeated measurements in a variety of locations on the same cell without destroying the integrity of the membrane. This flexibility has been especially useful to researchers for studying muscle cells as they contract under real physiological conditions, obtaining recordings quickly, and doing so without resorting to drastic measures to stop the muscle fibers from contracting.
A major disadvantage is that the resistance between the pipette and the membrane is greatly reduced, allowing current to leak through the seal, and significantly reducing the resolution of small currents. This leakage can be partially corrected for, however, which offers the opportunity to compare and contrast recordings made from different areas on the cell of interest. Given this, it has been estimated that the loose patch technique can resolve currents smaller than 1 mA/cm
2.
Patch-Seq
A combination of cellular imaging, RNA sequencing and patch clamp
this method is used to fully characterize neurons across multiple modalities.
As neural tissues are one of the most
transcriptomically diverse populations of
cells
Cell most often refers to:
* Cell (biology), the functional basic unit of life
Cell may also refer to:
Locations
* Monastic cell, a small room, hut, or cave in which a religious recluse lives, alternatively the small precursor of a monastery w ...
, classifying neurons into
cell types
A cell type is a classification used to identify cells that share morphological or phenotypical features. A multicellular organism may contain cells of a number of widely differing and specialized cell types, such as muscle cells and skin cells, ...
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
Electrophysiology (from Greek , ''ēlektron'', "amber" etymology of "electron"">Electron#Etymology">etymology of "electron" , ''physis'', "nature, origin"; and , '' -logia'') is the branch of physiology that studies the electrical properties of b ...
,
sequencing
In genetics and biochemistry, sequencing means to determine the primary structure (sometimes incorrectly called the primary sequence) of an unbranched biopolymer. Sequencing results in a symbolic linear depiction known as a sequence which succ ...
and
microscopy
Microscopy is the technical field of using microscopes to view objects and areas of objects that cannot be seen with the naked eye (objects that are not within the resolution range of the normal eye). There are three well-known branches of micr ...
, 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.
Automatic patch clamping
Automated patch clamp systems have been developed in order to collect large amounts of data inexpensively in a shorter period of time. Such systems typically include a single-use
microfluidic
Microfluidics refers to the behavior, precise control, and manipulation of fluids that are geometrically constrained to a small scale (typically sub-millimeter) at which surface forces dominate volumetric forces. It is a multidisciplinary field tha ...
device, either an
injection molded
Injection moulding (U.S. spelling: injection molding) is a manufacturing process for producing parts by injecting molten material into a mould, or mold. Injection moulding can be performed with a host of materials mainly including metals (for ...
or a
polydimethylsiloxane
Polydimethylsiloxane (PDMS), also known as dimethylpolysiloxane or dimethicone, belongs to a group of polymeric organosilicon compounds that are commonly referred to as silicones. PDMS is the most widely used silicon-based organic polymer, as its ...
(PDMS) cast chip, to capture a cell or cells, and an integrated electrode.
In one form of such an automated system, a pressure differential is used to force the cells being studied to be drawn towards the pipette opening until they form a gigaseal. Then, by briefly exposing the pipette tip to the atmosphere, the portion of the membrane protruding from the pipette bursts, and the membrane is now in the inside-out conformation, at the tip of the pipette. In a completely automated system, the pipette and the membrane patch can then be rapidly moved through a series of different test solutions, allowing different test compounds to be applied to the intracellular side of the membrane during recording.
See also
References
External links
Alternative images for patch clamp variations
{{DEFAULTSORT:Patch Clamp
Neurophysiology
Electrophysiology
Laboratory techniques