Methods for peptide analysis of sub-microliter biological samples using MALDI-TOF mass spectrometry

Biological systems are chemically complex systems which change in response to the environments. These changes could be normal function or indication of disease and can be monitored by analyzing the content of signaling molecules. Many methods had been developed to monitor and characterize these changes. The techniques for small volume samples analysis are limited. This thesis is to present the tools for sample handling and analysis methods to improve the detection of peptides from volume limited biological samples using MALDI-TOF-MS. A sub-microliter dialysis device was developed for removal of proteins from sub-microliter samples using a semi-permeable hollow fiber membrane to enhance small molecules detection by CE-LIF and peptide detection by MALDI-TOF-MS. To aid the analysis of complex peptide mass spectra obtained using MALDI-TOF, an automated Matlab algorithm was developed for finding significant peaks from mass spectra of healthy and diabetic patients. MALDI-TOF-MS method is also described to characterize peptide content of hemolymph collected from individual fruit fly larvae, which usually range 50-350 nL sample volume. Finally, an in-line nano-flow electroosmotic pump fabricated from fused-silica capillaries packed with silica gel is described for nanoliter sample delivery to assist in the automation of in vivo sample analysis. The author hopes the techniques presented in this thesis will assist and enable the analysis of volume-limited samples so that we can expand our research to smaller biological systems.