This dissertation describes the development and use of novel chemical affinity templates to direct the assembly and preserve the activity of biological molecules in surface- and solution-based biomolecular assays. The majority of work described herein focuses on advancing biological nanoarray technology to enable the study of fundamentally important biological processes that cannot currently be probed using the more conventional microfabrication approaches. Using high-resolution nanolithographic techniques, such as dip-pen nanolithography (DPN), nanoaffinity templates of alkanethiols can be easily fabricated on a gold surfaces with a resolution of 15 nm. Using DPN we systematically designed and demonstrated how single biomolecules (i.e. viruses) can be site-isolated and stratigically arranged on a surface with control over their orientation through chemical affinity templates based on divalent metal ion coordination chemistry. Although the direct coupling of biological entities to metal ion affinity templates works well for homogeneous soltions, an alternatve approach was developed to be more compatiable with heterogeneous soltions of biomolecules. This approach takes advantage of a particular set of proteins, antibodies, that exhibit high specificity and selectivity, which further allows for the retention of biological activity after a target molecule of interest has been captured.Using these functional biological nanoarrays, we have developed two novel cell-based assays to investigate virus-cell infectivity and protein-cell interactions at a level unatainable with current microarray-based techniques. A solution-based approach was also employed for the development of a simple and robust biodiagnostic assay for pathogenic microbes (viruses and bacteria). This approach takes advantage of oligonucleotide-functionalized gold nanoparticles, where the oligonucleotides can serve as recognition elements for a specific microbe. When these modified gold nanoparticles encounter a specific target molecule, they assemble and can easily and quickly be identified using their unique optical properties. Using this, we have developed a colorimetric and scanometric nanoparticle-based assay to detect and differentiate between three important human immunodeficiency virus subtypes. Finally, this disseration describes a real-time PCR based method, widly used in the biological and medical communities, to detect and quantify the number of oligonucleotides (barcodes) on a gold nanoparticle that is used as a target probe in a biodiagnostic assay (i.e. bio-barcode assay)

Last modified

• 06/05/2018

Creator

• Rafael Angel Vega

DOI

• https://doi.org/10.21985/N2CM4N

Subject

• Chemistry

Keyword

• biorecognition
• metal ions
• surface
• nanoarrays
• microscopy
• scanning
• probe
• viruses

Date created

• 2007-05-11

Resource type

• Dissertation

Rights statement

• In Copyright