EVALUATION OF A MICROWAVE-ASSISTED METAL-LABELING STRATEGY FOR QUANTIFICATION OF PEPTIDES VIA RPLC-ICP-MS AND THE METHOD OF STANDARD ADDITIONS
Steven J. Christopher, Eric L. Kilpatrick, Lee L. Yu, William C. Davis, blakely adair
NIST has performed preliminary research on applying a calibration methodology based on the method of standard additions to the quantification of lanthanide-labeled peptides via reverse-phase liquid chromatography coupled to inductively coupled plasma mass spectrometry (RPLC-ICP-MS). A microwave labeling procedure was developed and applied to derivatize peptides using the macrocyclic bifunctional chemical chelator DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), which significantly improved the lanthanide labeling yield and reduced reaction times relative to benchtop labeling procedures. Biomolecular MS technologies of matrix-assisted laser desorption ionization (MALDI)-MS and electrospray ionization (ESI)-MS/MS were used in concert with ICP-MS to confirm the results of microwave labeling, sample cleanup and standard additions experiments for several test peptides. The calibration scheme is outlined in detail and contextualized against complementary high accuracy calibration strategies currently employed for ICP-MS detection of biomolecules. Standard additions experiments using native, non-isotopic peptide calibrants confirm the simplicity of the scheme and the potential of applying a blending (recombined sample and spike) procedure, facilitating calibration via co-elution of lanthanide labeled peptides. Ways to improve and fully leverage the analytical methodology are highlighted.
peptide, standard additions, LC-ICP-MS, MALDI-MS, ESI-MS/MS, DOTA, Lanthanide