Direct vertical electroelution of protein from a PhastSystem band for mass spectrometric identification at the level of a few picomoles.
An electroelution apparatus prototype of a new design was constructed. In that design, the electric field passes vertically through the protein band located on a horizontal (PhastSystem) minigel polymerized on a net of Gel-Fix (Serva). A simple, home-made apparatus allows for electroelution of protein bands at the level of a few picomoles and their identification, after concentration, by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The technique is applicable to one-dimensional (1-D) or two-dimensional (2-D) gels of any size, but has been exemplified only by application to 1-D minigels to demonstrate the lower limits of protein load of the method. When in the course of further development of the prototype it will be combined with a modification to two dimensions of the electroelution mechanism under computer control of the high-performance gel electrophoresis apparatus (formerly of LabIntelligence), the new design appears uniquely qualified for an automated spot elution from 2-D gels under avoidance of gel sectioning.
Buzás
Z
Z
Section on Macromolecular Analysis, Laboratory of Cellular and Molecular Biophysics, NICHD, NIH, Bethesda, MD, USA.
Chang
H T
HT
Vieira
N E
NE
Yergey
A L
AL
Stastna
M
M
Chrambach
A
A
D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D008832:Microchemistry / Q000379:methods*; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D012710:Serum Albumin, Bovine