Electrophysiology

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A guide to Electrophysiology and Biophysics Laboratory Techniques

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Electrophysiology Lab Setup: Patch Clamp Rig

Electrophysiology is one of the foundational disciplines in neuroscience and cardiac physiology for the evaluation of ion channels. The patch-clamp technique is a versatile electrophysiological tool for understanding ion channel behavior.

Patch clamping has a broad range of applications in both basic research and drug discovery and development however, the complexities of the method can seem intimidating. To help researchers get up-to-speed, we’ve created this brief introduction to an electrophysiological lab patch clamp equipment setup.

 

Patch Clamp rig

Patch Clamp Equipment

 

What is it? The amplifier contains the circuitry required to measure electrical currents passing through ion channels or changes in cell membrane potential.

Why use it? Amplifiers measure changes in current or voltage.

The amplifier contains the circuitry necessary to measure current passing through the cell membrane both in magnitude and direction. The amplifier can also measure the cell membrane potential in response to the movement of current. The ideal place to amplify the signal is inside the recording instrument. All models of Axon™ amplifiers use this strategy with variable gain control on the output to provide low-noise amplification of the pipette current or membrane potential. Placing the amplification inside the recording instrument minimizes the amount of circuitry between the low-level signal and amplifying circuitry reducing extraneous noise sources.

Available amplifiers: Axopatch™ 200B, MultiClamp™ 700B, Axoclamp™ 900A

What is it? The digitizer is a data acquisition instrument that converts analog signals into digital signals.

Why use it? Digitizers capture data for analysis.

The current acquired by the amplifier is an analog signal, but in order to perform data analysis needed for high resolution patch-clamp measurements, the analog signal must be converted into a digital one. Positioned between the amplifier and the computer, the digitizer accomplishes this important task. The signal quality the computer receives is extraordinarily important, and this is determined by the sampling frequency, or sampling rate. The latest generation of Digidata® digitizers has the capability of sampling at 500 kHz and is equipped with the HumSilencer™ feature, which can eliminate 50/60 Hz line-frequency noise.

Available amplifiers: Digidata 1550B Low Noise Data Acquisition System plus HumSilencer

What is it? Patch clamp data and acquisition and analysis software is your interface with the amplifier, digitizer, and any other patch-clamp electronics.

Why use it? To perform data acquisition and data analysis, as well as to control the digitizer and amplifier.

While the amplifier and digitizer together hold the key circuitry that implements a patch-clamp experiment, the software controls these instruments so they deliver the desired potential(s) and measure the resulting current or voltage. In addition, the software analyses the acquired signal with user-defined settings, which can include filtering, normalization, noise removal, curve fitting, and parameter determination.

Available amplifiers: pCLAMP™ 11 Software

What is it? A device that holds the micropipettes with built-in circuitry to transmit electrical signals from the micropipettes onto the amplifier.

Why use it? The electrical signal acquired by the micropipette needs to be transmitted to amplifier systems for signal processing.

Each headstage is specifically tuned for the amplifier. All headstages contain critical electric circuitry that reduce noise. The headstage is also mechanically controlled by the micromanipulator.

Available headstages: Axon headstages

What is it? The microscope is an optical magnification tool. The micromanipulator is a device that mechanically maneuvers the micropipette with nanometer precision, typically allowing 3-dimensional movements.

Why use it? To precisely and stably position the micropipette to the area of cell membrane, which is critical for successful recording.

Accurately placing a patch electrode onto a 10-20 µm cell requires an optical system that can magnify up to 300- or 400- fold with contrast enhancement (e.g. Nomarski/DIC, Phase, or Hoffman) and a micromanipulator that stably positions the electrode in 3D space. An inverted microscope is preferable because it allows easier access for electrodes from above the preparation and also provides a larger, more solid platform to bolt the micromanipulator. A micromanipulator has the ability to move the electrode in very minute distances along the X, Y, and Z axes. The micromanipulator can then hold that position indefinitely.

What is it? A table and cage around your patch-clamp setup to isolate sources of interference.

Why use it? To shield your setup from external interference.

Electrical currents measured during patch-clamp experiments can be extremely small (in the pico-amp range), and any small sources of interference, such as radio waves, can distort or obscure these signals. A Faraday cage is a wire mesh enclosure around your microscope and recording chamber; it is useful in preventing the electrodes from picking up extraneous noise sources. Additionally, small sources of vibration on the order of pico-meter magnitude can disrupt your recording. Hence, all components must be perfectly positioned throughout the time-course of your experiment, and the air or anti-vibration tables are used to isolate your setup from external sources of vibration that may disrupt this alignment.

Webinar

Save your time on data analysis with new Batch Analysis feature in Axon pCLAMP 11 Software

Speaker: Jeffrey Tang, Ph.D.

Senior Global Axon Electrophysiology Application Scientist

Tuesday, March 24, 2020

10:00 AM PDT / 1:00 PM EDT

Join our webinar and learn how to use the Clampfit software module to manipulate and analyze electrophysiological data acquired by Axon pCLAMP™ 11 Software. The software’s advanced Batch Analysis macros eliminate the need to define parameters for every set, streamlining data analysis. Dr. Jeffrey Tang will provide an overview of the new Batch Analysis features, demonstrate the use of macros, analyze batch data, and plot graphs.

Register now

 

Patch Clamping Technology

The Axon™ instrument portfolio provides a comprehensive solution for patch-clamping that includes amplifiers, digitizer, software, and accessories. Our best-in-class instruments facilitate the entire range of patch-clamp electrophysiology techniques from the smallest single-channel to the largest macroscopic recordings. 

The Axon pCLAMP™ 11 Software Suite is the most widely used electrophysiology data acquisition and analysis program for control and recording of voltage-clamp, current-clamp, and patch-clamp experiments. Several key features listed below help streamline workflow, allowing you to do more sophisticated experiments, execute them more effciently and generate higher quality data.

 

  • Automated Event Detection

    Clampfit Automated Event Detection

    Clampfit Advanced Analysis Module, part of the pCLAMP 11 Software suite, has a flexible event detection engine that analyzes spontaneous and evoked action potentials and post-synaptic data. Events are detected by threshold crossing or a pattern-matching Template Search. Template Searches analyze spontaneous events like miniature synaptic EPSPs and IPSPs. Additionally, multiple event categories of events can be simultaneously detected. The integrated environment of Clampfit 11 Software links detected events in data to the spreadsheet and graph windows enabling quick contextual evaluation of the entire dataset.

    Batch Data Analysis Macros

    Batch Data Analysis Macros

    Clampfit Advanced Analysis Module, part of the pCLAMP 11 Software suite contains a Batch Data Analysis Tool that utilizes macros to accelerate data analysis. Batch analysis saves time by analyzing abundant amounts of data created by the same protocol. To use batch analysis, simply turn on the macro capture feature, analyze the data, and save the macro. When additional data needs analyzing, simply apply the saved macro and the data is analyzed automatically.

  • Population Spike Search Tool

    Population spike recordings and paired-pulse experiments, while simple to collect, have traditionally been difficult to analyze. That is no longer the case with the Clampfit Advanced Analysis Module in pCLAMP 11 Software. The Population Spike Search tool will automatically locate population spikes based on user defined parameters and calculate the amplitude, area under the curve, half-width, rise time, decay time, rise slope, decay slope, and coastline of population spikes and paired-pulses.

    HumSilencer Technology

    HumSilencer Technology

    50/60 Hz line-frequency noise, also known as electrical hum, is the most common source of background noise in patch-clamp electrophysiology experiments. This noise can overwhelm biological signals of interest, making sensitive patch-clamp measurements nearly impossible. Traditional trouble shooting is typically only partially effective and can impair data accuracy. HumSilencer is a filter-free, adaptive technology that learns and removes line frequency noise without using methods that impair signal accuracy like filters, that can distort biological signals.

    Read more  

  • Action Potential Search

    Patch-Clamp Action Potential Search for Cellular Events

    Action potentials represent important cellular events. Without action potentials, hearts would not beat, and neurons would not fire, so measuring these events is essential. The Action Potential Search tool in Clampfit 11 Advanced module detects all action potentials in the data file. It applies user defined and programmatically determined metrics including amplitude, AP duration, rise and decay time, rise and decay slope, peak to peak frequency and time, amplitude delta per peak, afterpotential amplitude and duration, and threshold potential.

    The Axon Guide

    The Axon Guide

    A guide to Electrophysiology and Biophysics Laboratory Techniques. The purpose of this guide is to serve as an information and data resource for electrophysiologists. It covers a broad scope of topics ranging from the biological basis of bioelectricity and a description of the basic experimental setup to a discussion of mechanisms of noise and data analysis.

    Download Guide  

Resources of Electrophysiology

Videos and Demos

Population Spike Analysis in pCLAMP 11

Population Spike Analysis in pCLAMP 11

Molecular Devices Batch Data Analysis Macros

Batch Data Analysis Macros

Action Potential Analysis in Clampfit Module

Action Potential Analysis in Clampfit Module

Related Products & Services of Electrophysiology