Bas Jansen

The research in this thesis can be divided into two parts, i.e. method development and method application, which are both introduced in Chapter 1. The method development part focusses on the targeted quantitation of glycans and glycopeptides, while the method application part uses the developed methods to study glycosylation during pregnancy and the differences between maternal and fetal serum. This section presents a of the main findings of each chapter of the thesis.

Chapter 2 of the thesis describes the development of a software package for the targeted quantitation of glycans and glycopeptides measured by matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS). The aim was to develop a tool that would allow high-throughput preprocessing of MALDI-TOF-MS data, i.e. calibration of mass spectra, peak area integration and the calculation of a variety of quality control parameters. There was an obvious need for such a tool as the existing software packages only were capable of fulfilling parts of the desired functionality or were not meant for high-throughput use. The developed tool “MassyTools” is a graphical software package that performs calibration and quantitation of analytes on the basis of a user-defined list of analytes. Furthermore, MassyTools is capable of calculating several quality control parameters, e.g. mass accuracy and isotopic pattern quality, thereby allowing for straightforward spectrum and analyte curation. The results showed that MassyTools yielded a higher quality calibration, in the form of lower mass errors post calibration, than the most commonly used commercial software package. Furthermore, the quantitation of low-abundant analytes using MassyTools also provided a higher precision.

Chapter 3 of the thesis describes the development of LaCyTools, a targeted software package for glycans and glycopeptides measured by liquid chromatography(LC)-electrospray ionization (ESI)-MS. The alignment of LC-MS runs is based on a user-defined list of features. The quantitation is performed using a sum spectrum per glycopeptides cluster as the glycoforms on a single glycosylated peptide tend to elute at a similar retention time in our system. A major difference between MALDI-TOF-MS and LC-ESI-MS data is the presence of multiple charge states in LC-ESI-MS. Therefore, LaCyTools was designed to handle multiple charge states in parallel with the user specifying the minimum and maximum charge states. LaCyTools was compared with several software packages, showing unparalleled throughput as more than 200 LC-MS datasets could be processed within 10 hours. Furthermore, LaCyTools offered precise quantitation