Whole-cell patch-clamp: true perforated or spontaneous conventional recordings?
Perforated whole-cell patch-clamp recordings obtained with nystatin are frequently used to preserve intracellular integrity. However, the perforated-patch configuration may sometimes undergo a spontaneous change into the conventional whole-cell configuration, especially when lymphocytes are investigated. The electrophysiological criteria-- previously described--for establishing the existence of the perforated whole-cell configuration have been shown to be insufficient. Thus, the dye eosin, applied to the pipette solution, was tested as a tool for discriminating between the perforated and the conventional whole-cell configurations on rat T-lymphocytes. The dye never entered the cell from the pipette during the entire measurement in the perforated whole-cell configuration. In contrast, all cells in the conventional whole-cell configuration became red immediately after membrane rupture. Eosin barely changed the currents studied. The results suggest that eosin is a dye of choice for verifying a true perforated-patch configuration.
Strauss
U
U
Department of Neurology, University of Rostock, Germany. ulf.strauss@med.uni-rostock.de
Herbrik
M
M
Mix
E
E
Schubert
R
R
Rolfs
A
A
D000818:Animals; D002460:Cell Line; D004801:Eosine Yellowish-(YS) / Q000493:pharmacokinetics; D005456:Fluorescent Dyes / Q000493:pharmacokinetics; D007476:Ionophores / Q000494:pharmacology; D051664:Kv1.3 Potassium Channel; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009761:Nystatin / Q000494:pharmacology; D018408:Patch-Clamp Techniques / Q000379:methods* / Q000592:standards; D015221:Potassium Channels / Q000502:physiology; D024642:Potassium Channels, Voltage-Gated; D051381:Rats; D011917:Rats, Inbred Lew; D015203:Reproducibility of Results; D013601:T-Lymphocytes / Q000166:cytology / Q000502:physiology*
D005456:Fluorescent Dyes; D007476:Ionophores; C496975:Kcna3 protein, rat; D051664:Kv1.3 Potassium Channel; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D009761:Nystatin; D004801:Eosine Yellowish-(YS)