Analysis and localization of metal‐ and metalloid‐containing proteins by synchrotron radiation x‐ray fluorescence spectrometry

Abstract

AbstractSynchrotron radiation x‐ray fluorescence (SRXRF) spectrometry has been applied for the determination of metals and metalloids in electrophoretically separated proteins and histological samples. The method was tested by the analysis of trace element‐containing proteins in rat testis homogenate and the determination of the spatial distribution of some metals in brain sections of scrapie‐infected hamsters. After sodium dodecyl sulfate—polyacrylamide gel electrophoresis (SDS‐PAGE) and transfer on to a blotting membrane, the distribution of selenium among the proteins of testis homogenate was determined. The comparison with the autoradiogram of 75Se‐labeled testicular proteins separated in the same way showed that of nine selenium‐containing proteins found by autoradiography, four could be detected by SRXRF. The distribution of several metals and metalloids in cryo‐sections of 10 µm thin brain sections of three Syrian hamsters intracerebrally infected with the scrapie strain ME7‐H and of three control animals was investigated at the HASYLAB beamline L by scanning with a synchrotron radiation beam focused by a polycapillary lens. The copper‐binding prion protein (PrP) in its pathological form PrPSc, determined by immunohistochemistry on cryo‐sections, was found to be diffusely dispersed in the scrapie‐infected brains, and the copper distribution differed from that in the control animals. The question whether, and to what extent, PrPSc and copper hot‐spots are related, remains to be investigated. The method was shown to be suitable for the determination of trace element distribution patterns in electrophoretically separated proteins and histological tissue preparations. Copyright © 2004 John Wiley & Sons, Ltd.