The primary objective of the MSC Electrophysiology Core is to provide a service and expertise to researchers who plan to conduct experiments requiring electrophysiological techniques, such as patch-clamp electrophysiology to determine the functions of ion channels, transporters or receptors, or the electrical activities of neurons or muscles.
Patch-clamp electrophysiology and its applications
Patch-clamp electrophysiology is the study of electrical activity across biological membranes such as the plasma, mitochondrial and lysosomal membrane. Movement of ions such as Na+, K+, Ca2+ or Cl- or other charged molecules, across the membrane, can in principle be monitored via recordings as current or membrane potential change. The technique can therefore be used to determine and record membrane excitability of neurons or muscles in terms of action potential firings or amplitudes of ionic currents. Besides using the technique on muscles and neurons, the method has been applied to measure ion flux through pore-forming proteins coded by genes of the SARS and influenza viruses or through ectopic expression of ion channels in cancer cells. In essence, the technique can be used for functional study by monitoring charge movements mediated by a specific protein or by your protein of interest that is found on the plasma membrane.
To perform whole cell patch-clamp, a micropipette is tightly sealed on the cell membrane forming high resistance gigaseal. This small patch of membrane could then be ruptured with more intense suction to allow for electrode electrical access to the interior of the cell. Voltage stimulus is subsequently applied to elicit membrane current. Alternatively, change in membrane potential, for example action potential which is an important feature of excitable cells such as neurons or muscle fibers, could also be recorded.
Key capabilities of the MSC Electrophysiology Core:
- Whole cell recording of cultured cells including:
- Functional characterisation of ion channels in transfected cell lines
- Pharmacological screening of ion channel blockers/activators
- Functional characterisation of human iPSC derived neurons or cardiomyocytes
- Whole cell recording of isolated cardiomyocytes or cultured primary neurons
- Whole cell recording of neurons within acutely isolated brain slice
Workflow of Electrophysiology Core:
The Facility will purchase to provide for the general consumables and reagents that will be proportionately charged to the users. However, users are responsible to supply any materials or reagents that are unique to their particular projects.
|Types of service||Charges for NUS/NUHS staff||Charges for others||Description|
|Culture and transfection of HEK293 cells for patch clamp||S$120.00||S$180.00||
Includes coating of cover slips with PDL, splitting of cells and transfection with lipofectamine 2000 sufficient for two days of recording
|Preparation of brain slice||S$30.00||S$45.00|
|Isolation of mouse cardiomyocytes||S$80.00||S$120.00|
|Charges for NUS/NUHS staff (per hour)||Charges for others
|Whole cell patch clamp electrophysiology of transfected HEK293 cells||S$60.00||S$90.00||
Includes whole-cell patch clamp analysis and preparation of internal and external solution
|Whole cell patch clamp electrophysiology of brain slice||S$60.00||S$90.00|
|Whole cell patch clamp electrophysiology of cardiomyocytes||S$60.00||S$90.00|
|Data analysis and figures preparation by the electrophysiology core (experimental design, data acquisition, analysis and interpretation).||Request for authorship contribution||Request for authorship contribution|
Prices quoted are subject to changes.
Huang Hua, PhD
Tel no: (65) 6516 8166; Fax No: (65) 6777 3271