ISOTOPE-DILUTION MASS-SPECTROMETRIC QUANTIFICATION OF SPECIFIC PROTEINS - MODEL APPLICATION WITH APOLIPOPROTEIN-A-I

An enzymatic hydrolysis isotope dilution-mass spectrometric method was developed for reference quantification of specific proteins. The anal ytical procedure involved measuring a reproducibly hydrolyzed peptide (serving as the primary standard) unique to a specific protein, This n ew mass spectrometric method was evaluated by assessing the concentrat ion of apolipoprotein (apo), A-I in the European Community Bureau of R eference (BCR) lyophilized Certified Reference Material (CRM 393). We used the method to make 96 measurements (4 replicate analyses of 4 enz ymatic digests of 6 vials of BCR-CRM 393), which gave an average total protein mass of 1.048 mg (+/- 1.0% at 99% confidence limits). The tot al overall analytical CV was 3.95%. The results of this evaluation of our model approach to determine the concentration of a specific protei n in a purified preparation demonstrated that our new mass spectrometr ic method can be used to measure apolipoproteins and other specific pr oteins without the use of epitopic immunoassay methods.