Matrix Assisted Laser Desorption / Ionization (MALDI) Mass Spectrometry (MS) has emerged as an important tool in the field of proteomics mainly because it is simple, quick and efficient. The identification and quantitation of biomarkers, protein targets for drugs, and metabolites are some of the important fields in proteomics research. Although MALDI MS is an important tool in proteomics research there are drawbacks of the technique that need further development in order for the approach to be used in clinical laboratories. One major limitation of MALDI MS is the generally poor reproducibility of ion signal intensities, which negatively impacts the quantitation of peptides and protein by MALDI MS. A considerable amount of research has been performed in an effort to improve the ion signal reproducibility in MALDI MS. However, many of the approaches developed have introduced specific drawbacks with respect to the traditional dried-droplet sample preparation technique, negating many of the advantages of the MALDI MS approach. This project has focused on the development of a novel approach to quantify peptides by MALDI MS while preserving traditional known advantages of the technique. The studies performed show that an approach in which the ion signal base widths are manipulated to match that of a reference ion signal, through adjustments in desorption laser intensity, leads to much higher reproducibility in the integrated ion signal intensities. A standard curve acquired using the constant ion signal base width approach showed lower average RSDs (< 10.00% vs.> 39.00%) and improved R2 values (> 0.9600 vs. < 0.809) as compared to the conventional constant desorption laser intensity approach. Subsequent work also revealed that the peptide hydrophobic / hydrophilic properties influenced the applicability of the quantitation approach to mixtures of peptides. Specifically, the data revealed that peptides with differing hydrophobic / hydrophilic properties appear to co-crystallize with the MALDI matrix differently leading to an inability to use a hydrophobic peptide signal to quantitate a hydrophilic peptide, and vice versa. This latter conclusion was further supported in similar studies performed on the mixture of peptides resulting from tryptic digestion of the protein bovine serum albumin.
| Identifer | oai:union.ndltd.org:siu.edu/oai:opensiuc.lib.siu.edu:dissertations-1993 |
| Date | 01 May 2015 |
| Creators | Priyasantha, Kandalama KD |
| Publisher | OpenSIUC |
| Source Sets | Southern Illinois University Carbondale |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Dissertations |
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