TY - JOUR
T1 - Isolating nuclei from cultured cells for patch-clamp electrophysiology of intracellular Ca(2+) channels
AU - Mak, Don On Daniel
AU - Cheung, King Ho
AU - Vais, Horia
AU - Foskett, J. Kevin
PY - 2013/9/1
Y1 - 2013/9/1
N2 - Nuclear patch-clamp experiments can be performed with intact nuclei or with nuclei from which the outer nuclear membrane has been removed. This protocol presents procedures for harvesting different types of cultured cells, isolating nuclei, and exposing the inner nuclear membrane by agitating in the presence of sodium citrate. Particulars about obtaining and maintaining the cells of interest in culture are not described here. However, care should be taken not to allow the cells to grow beyond a density of 2–3 × 106 cells/mL because this may decrease both the cell viability and the success rate of detecting active inositol 1,4,5-trisphosphate receptor (InsP3R) channels in nuclear patches.
AB - Nuclear patch-clamp experiments can be performed with intact nuclei or with nuclei from which the outer nuclear membrane has been removed. This protocol presents procedures for harvesting different types of cultured cells, isolating nuclei, and exposing the inner nuclear membrane by agitating in the presence of sodium citrate. Particulars about obtaining and maintaining the cells of interest in culture are not described here. However, care should be taken not to allow the cells to grow beyond a density of 2–3 × 106 cells/mL because this may decrease both the cell viability and the success rate of detecting active inositol 1,4,5-trisphosphate receptor (InsP3R) channels in nuclear patches.
UR - http://cshprotocols.cshlp.org/content/2013/9/pdb.prot073056.abstract?sid=c9e5d07d-ed60-4b87-8113-0e7516f058b8
UR - https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3979423/
M3 - Journal article
SN - 1940-3402
VL - 9
SP - 880
EP - 884
JO - Cold Spring Harbor Protocols
JF - Cold Spring Harbor Protocols
ER -