Speciation of metal ions in proteins by combining PIXE and thin layer electrophoresis

Particle induced X-ray emission (PIXE) spectroscopy is a simple and convenient method of quantitative multielemental analysis with sensitivities in the μg/g range, that can be successfully used for trace analysis of metal ions in proteins or enzymes. However, due to its elemental character the technique alone is not a priori suitable for speciation. Keeping track of the metal ions of interest throughout a proper biochemical separation technique, on the other hand, could be a useful strategy for speciation. Different versions of thin layer electrophoresis (polyacrylamide gel, agarose or cellulose acetate electrophoresis) are very effective and sensitive methods to separate proteins or protein fragments. Due to the high absolute sensitivity of PIXE the metal ions concentrated in the narrow bands of an electropherogram can be in situ successfully detected. The present paper describes this unique combination of biochemical separation and ion beam analysis which significantly extends the information obtained from electrophoresis. Illustrative applications are given and the advantages and limitations of the method are discussed. Possible extensions of the technique are also outlined.