Direct vertical electroelution of protein from a PhastSystem band for mass spectrometric identification at the level of a few picomoles

• Published in: Proteomics (Weinheim) 2001-05, Vol.1 (5), p.691-698
• Main Authors: Buzás, Zsuzsanna, Chang, Huan-Tsung, Vieira, Nancy E., Yergey, Alfred L., Stastna, Miroslava, Chrambach, Andreas
• Format: Article
• Language: English
• Subjects: electroelution; Electrophoresis, Polyacrylamide Gel - methods; Mass spectrometry; Microchemistry - methods; Picomolar protein identification; protein purification; proteomics; Serum Albumin, Bovine - chemistry; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods; Vertical electroelution
• ISSN: 1615-9853; 1615-9861
• DOI: 10.1002/1615-9861(200104)1:5<691::AID-PROT691>3.0.CO;2-7

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.