Abstract:
Medical diagnosis is the process of attempting to determine and/or identify a
possible disease or disorder. This process is revealed by biomarkers, defined
by The Food and Drug Administration (FDA) as “characteristics that are
objectively measured and evaluated as indicators of normal biologic
processes, pathogenic processes, or pharmacologic responses to a
therapeutic intervention”. The process of biomarker discovery has been
boosted in the last years by proteomics, a research discipline that takes a
snapshot of the entire wealth of proteins in an organism/ tissue/ cell/ body
fluid. An implementation of the analysis methods can help in isolate proteins
present in the low range of concentrations, such as biomarkers very often
are. An established biomarker can further be measured with the help of
biosensors, devices that can be employed in the point-of care diagnostics.
This PhD thesis shows and discusses the results of three projects in the field
of protein biomarkers discovery and quantification. The first project exploited
proteomics techniques to find relevant protein markers for Intrauterine
Growth Restriction (IUGR) in cordonal blood serum (UCS) and amniotic
fluid (AF). A 14 proteins in UCS and 11 in AF were successfully identified and
found to be differentially expressed.
Molecularly Imprinted Polymers (MIPs) directed towards proteins and
peptides containing phosphotyrosine were then produced, with the final goal
of selectively extracting phosphopeptides from a peptide mixture. An
alteration of the phosphorylation pattern is in fact often associated to
important diseases such as cancer. The polymers were produced as
nanoparticles, that were characterised with Dynamic Light Scattering (DLS)
and Atomic Force Microscopy (AFM). A recipe was also tested for binding
capacity towards phosphotyrosine.
A Surface Plasmon Resonance (SPR) biosensor to quantify hepcidin
hormone was finally produced. This is the major subject in iron homeostasis
in vertebrates and marker of iron unbalance diseases. A calibration curve
was made and affinity/kinetic parameters for the ligand employed were
measured.
