Fully automated two-dimensional electrophoresis system for high-throughput protein analysis.
We developed a fully automated electrophoresis system for rapid and highly reproducible protein analysis. All the two-dimensional (2D) electrophoresis procedures including isoelectric focusing (IEF), on-part protein staining, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and in situ protein detection were automatically completed. The system comprised Peltiert devices, high-voltage generating devices, electrodes, and three disposable polymethylmethacrylate (PMMA) parts for IEF, reaction chambers, and SDS-PAGE. Because of miniaturization of the IEF part, rapid IEF was achieved in 30 min. A gel with a tapered edge gel on the SDS-PAGE part realized a connection between the parts without use of a gluing material. A biaxial conveyer was employed for the part relocation, sample introduction, and washing processes to realize a low-maintenance and cost-effective automation system. Performances of the system and a commercial minigel system were compared in terms of detected number, resolution, and reproducibility of the protein spots. The system achieved high-resolution comparable to the minigel system despite shorter focusing time and smaller part dimensions. The resulting reproducibility was better or comparable to the performance of the minigel system. Complete 2D separation was achieved within 1.5 h. The system is practical, portable, and has automation capabilities.
Hiratsuka
Atsunori
A
Research Center of Advanced Bionics, National Institute of Advanced Industrial Science and Technology, c/o Katayanagi Institute, Tokyo University of Technology, 1404-1, Katakura, Hachioji, Tokyo 192-0982, Japan.
Kinoshita
Hideki
H
Maruo
Yuji
Y
Takahashi
Katsuyoshi
K
Akutsu
Satonari
S
Hayashida
Chie
C
Sakairi
Koji
K
Usui
Keisuke
K
Shiseki
Kisho
K
Inamochi
Hajime
H
Nakada
Yoshiko
Y
Yodoya
Kouhei
K
Namatame
Ichiji
I
Unuma
Yutaka
Y
Nakamura
Makoto
M
Ueyama
Kosuke
K
Ishii
Yoshinori
Y
Yano
Kazuyoshi
K
Yokoyama
Kenji
K
D001331:Automation; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D007525:Isoelectric Focusing / Q000379:methods; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D015203:Reproducibility of Results; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013194:Staining and Labeling
D011506:Proteins; D012967:Sodium Dodecyl Sulfate