G-electrode-loading method for isoelectric focusing, enabling separation of low-abundance and high-molecular-mass proteins.
The G-electrode-loading method (GELM) is a technique enabling a large number of proteins from rat liver to enter an immobilized pH gradient (IPG) gel strip for isoelectric focusing (IEF). In this method, three slips containing the sample solution are placed on the cathodic edge of an IPG gel strip and a slip containing Chaps solution, a filtration membrane, and an electrode slip are placed on top. Finally, a G-electrode is placed on these slips. The Chaps solution (an amphoteric compound) is supplied gently to the sample solution during IEF and helps the proteins in the sample solution to enter the IPG gel strips with a high solubilization capacity. This method was compared with traditional slip-loading and in-gel rehydration, and it showed the best results for protein separation, including high-molecular-mass proteins.
Koga
Kaori
K
Anatech Corporation, Bunkyo, Tokyo, Japan. kdobashi@anatech.co.jp
D000818:Animals; D004566:Electrodes; D015180:Electrophoresis, Gel, Two-Dimensional; D007525:Isoelectric Focusing / Q000379:methods*; D008970:Molecular Weight; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D018892:Proton-Motive Force; D051381:Rats
D011506:Proteins