Rapid extraction and structural characterization of biomolecules in agarose gels by laser desorption Fourier transform mass spectrometry.
A method originally developed for the extraction of biomolecules from agarose gel slices has been utilized as a rapid means of isolating biological compounds from gels for subsequent structural characterization by matrix-assisted last desorption-ionization Fourier transform mass spectrometry (MALDI/FTMS). This "freeze-squeeze" extraction method involves pressure extrusion of fluid from frozen gel slices and provides near 50% recovery of analyte in less than 5 min. Experiments were directed at examining the recovery efficiency of the extraction method using 14C-labeled adenosine monophosphate and investigating the effect of high buffer concentrations on the laser desorption mass spectra. When coupled with this extraction technique, MALDI/FTMS can be used to detect and identify biomolecules at the low picomole level in agarose gel slices. The accurate mass measurements and MS/MS capabilities of the FTMS were exploited to provide detailed structural information at the isomeric level for oligonucleotides electrophoresed into agarose gels.
Dunphy
J C
JC
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta 30332-0400.
Busch
K L
KL
Buchanan
M V
MV
D000249:Adenosine Monophosphate / Q000737:chemistry / Q000302:isolation & purification*; D004587:Electrophoresis, Agar Gel; D005583:Fourier Analysis; D013058:Mass Spectrometry; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000302:isolation & purification*
D009838:Oligodeoxyribonucleotides; D000249:Adenosine Monophosphate