Comparison of analytical methods using enzymatic activity, immunoaffinity and selenium-specific mass spectrometric detection for the quantitation of glutathione peroxidase 1

Abstract

Glutathione peroxidase 1 (Gpx1), one of the most responsive selenoproteins to the variation of selenium concentration, is often used to evaluate "selenium status" at a cellular or organismal level. The four major types of analytical methodologies to quantify Gpx1 were revisited. They include (i) an enzymatic assay, (ii, iii) polyacrylamide gel electrophoresis (PAGE) with (ii) western blot detection of protein or (iii) inductively coupled plasma mass spectrometry (ICP MS) detection of selenium, and (iv) size-exclusion chromatography with ICP MS detection. Each of the four methods was optimized for the quantification of Gpx1 with maximum sensitivity. The methods based on the enzymatic and immunodetection offer a much higher sensitivity but their accuracy is compromised by the limited selectivity and limited dynamic range. The advantages, drawbacks and sources of error of each technique are critically discussed and the need for the cross-validation of the results using the different techniques to assure the quality assurance of quantitative analysis is emphasized.

Keywords: Enzymatic assay; Gel electrophoresis; Glutathione peroxidase 1; ICP MS; LC; Laser ablation ICP MS; Selenium; Western blot.