| Popis: |
Understanding the interactions of biological molecules with solid supports is vital for the development of detection systems and assay platforms. These relationships are frequently quite complex, involving hydrophobic interactions, electrostatic interactions, van der Waals forces, and covalent chemical bonds. We can exploit these interactions in a solid support device by modifying the surface substrate with thin films and monolayers. Self-assembled monolayers (SAMs) are powerful tools for functionalizing and imparting chemical character to surfaces. In this thesis, alkylthiol reagents are utilized to build SAMs on gold (Au) substrates. This work characterizes and studies monolayer formation. In addition, I use SAMs to generate surfaces specific for binding proteins, DNA, and cells. The popular biotin-streptavidin motif is used to demonstrate protein binding, as well as characterize monolayer composition as a result of solvent effects. Novel reagent syntheses are presented for both biotinylated alkylthiols and triethylene-glycol alkylthiols. Together, these two reagents generate substrates which bind specific proteins, while repelling non-specific ones. An additional reagent, “DMT-coated controlled porous glass (CPG),” was designed and synthesized for the generation of custom sequence oligonucleotides. Phosphoramidite syntheses using this modified CPG yield oligos with a 3’ alkylthiol modification. SAMs generated with this reagent demonstrate specific binding of complement strands. Both electrochemical techniques and restriction enzymes (where appropriate) provide methods for releasing monolayer-bound species. Lastly, I employ SAMs to generate substrates amenable to cell capture and cell adhesion. Binding B- and T-cell lymphocytes is achieved, demonstrating SAM-coated Au as a substrate for cell panning. Chemokine vascular endothelial growth factor (VEGF) is also bound to SAMs, generating surfaces amenable to cell adhesion and motility. Cells plated on higher surface concentrations of VEGF migrate faster, and I show the effect is specific to cells with VEGF receptors. Overall, this thesis explores the formation and utilization of SAMs for capturing and studying biological targets. The findings here may be transferred in the future into bio-sensing devices and arrays. |
