Article Text

PDF
ASSA13-03-21 Establishment of Method For High-Throughput K+ Outward Current Recording on Jurkat Cell by Automated Patch Clamp Technology
  1. Huang Wenjun,
  2. Zhou Rui,
  3. Li Tao,
  4. Mao Liang,
  5. Zeng Xiaorong,
  6. Yang Yan
  1. Key Laboratory Of Medical Electrophysiology of Ministry of Education, Luzhou Medical College

Abstract

Objective Due to high-throughput capabilities, automated patch clamp system has been widely used in cardiovascular ion channel research in rescent years. The aim of this study is to establish the method for high-throughput K+ outward current recording on jurkat cell using automated patch clamp system.

Methods Using NPC-16 chip by which 8 cells could be recorded simultaneously, whole cell K+ outward current on Jurkat cell was recorded. Besides, inhibitory effects of Tetraethylammonium chloride (TEA) were monitored and IC50 for TEA was also determined. Optimization of cell culture condition and seal enhancer solution containing different Ca2+ concentrations (10, 20, 30, 35, 40, 45 mM respectively) was performed to improve success rate of sealing and whole-cell access.

Results For 96 cells from NPC-16 chips, the success rate of giga-seal and whole-cell access was 95% and 87% respectively. For cell culture optimization, efficient cell passaging by dissociating cell clusters to single cells could facilitate seal and whole-cell access. Among different concentration of Ca2+, 30 mM Ca2+ was found to be most efficient for giga-seal and whole-cell access. Moreover, The K+ outward current could be inhibited by TEA in a dose dependent manner with IC50 (10.6 ± 2.7 mM).

Conclusions Taken together, one method for high-throughput K+ outward current recording on Jurkat cell by automated patch clamp system was successfully established with high success rate of giga-seals and whole-cell access. Optimization of cell culture condition and seal enhancer solution could promote higher success rate. The system will allows for a cost-effective and high throughout cell currents recording in future cardiovascular disease research.

Statistics from Altmetric.com

Request permissions

If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.