NDLI: Minimizing Cytosol Dilution in Whole-Cell Patch-Clamp Experiments

Content Provider	IEEE Xplore Digital Library
Author	Inayat, S. Pinto, L.H. Troy, J.B.
Copyright Year	1964
Abstract	During a conventional whole-cell patch clamp experiment, diffusible cytosolic ions or molecules absent in the pipette solution can become diluted by a factor of one million or more, leading to diminished current or fluorescent signals. Existing methods to prevent or limit cytosol diffusion include reducing the diameter of the pipette's orifice, adding cytosolic extract or physiological entities to the pipette solution, and using the perforated patch clamp configuration. The first method introduces measurement error in recorded signals from increased series resistance and the latter two are cumbersome to perform. In addition, most perforated patch configurations, prevent investigators from using test compounds in the pipette solution. We present a method to overcome these limitations by minimizing cytosol dilution using a novel pipette holder. Cell-attached configuration is obtained with the pipette filled with pipette solution. Most of the pipette solution is then replaced with mineral oil so that cytosol dilution can be minimized in whole-cell configuration. To accomplish this requires a suction line and two Ag/AgCl electrodes inside the pipette. Testing our novel pipette holder with Chinese Hamster Ovarian cells, we demonstrate cytosol dilution factors between 76 and 234. For large cells with somas greater than 40 μm, cytosol dilution factors of 10 or less are achievable.
Sponsorship	IEEE Engineering in Medicine and Biology Society
Page Count	10
File Size	1222731
Starting Page	2042
Ending Page	2051
File Format	PDF
ISSN	00189294
Volume Number	60
Issue Number	7
Language	English
Publisher	Institute of Electrical and Electronics Engineers, Inc. (IEEE)
Publisher Date	2013-01-01
Publisher Place	U.S.A.
Access Restriction	One Nation One Subscription (ONOS)
Rights Holder	Institute of Electrical and Electronics Engineers, Inc. (IEEE)
Subject Keyword	Electrodes Wires Electron tubes Assembly Minerals Cells (biology) Polyimides whole cell Cytosol electrophysiology patch clamp pipette holder rundown
Content Type	Text
Resource Type	Article
Subject	Biomedical Engineering

Sl.	Authority	Responsibilities	Communication Details
1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
5	Website/Portal (Helpdesk)	Queries regarding NDLI and its services	support@ndl.gov.in
6	Contents and Copyright Issues	Queries related to content curation and copyright issues	content@ndl.gov.in
7	National Digital Library of India Club (NDLI Club)	Queries related to NDLI Club formation, support, user awareness program, seminar/symposium, collaboration, social media, promotion, and outreach	clubsupport@ndl.gov.in
8	Digital Preservation Centre (DPC)	Assistance with digitizing and archiving copyright-free printed books	dpc@ndl.gov.in
9	IDR Setup or Support	Queries related to establishment and support of Institutional Digital Repository (IDR) and IDR workshops	idr@ndl.gov.in

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