Development of new fluorescent molecules for use as biomolecular probes

This thesis is composed of two aspects of research that will be linked as part of a future PhD project. The first part describes the development of two new ligands designed to chelate lanthanide ions - specifically europium and terbium. The ligands are based on the known terphenyl platform (and are related to BHHCT), but replace the central phenyl ring with a 2,3-disubstituted quinoxaline ring. The goal of this aspect of the research is to prepare compounds capable of producing long-lived fluorescence that is either red (for Eu3+ chelates) or green (for Tb3+ chelates) that can be conjugated to biomolecules to form biological probes. The second part is aimed at preparing a fluorescent peptide conjugate from a linker peptide (LPG) with conventional fluorescent dyes from the Alexa Fluor family. The ability of this conjugate to directly label an antibody to quickly and efficiently generate a fluorescent antibody for use as a biomolecular probe was also investigated. Utimately, the new lanthanide chelates described in the first part of this work will be covalently bound to Linker Protein G (LPG). LPG is a recombinant fusion protein consisting of two regions; (a) a peptide linker sequence containing eight lysine residues and (b), Streptococcus Protein G' which has specific binding affinity towards antibodies. The lanthanide chelates will be attached to LPG via the lysine residues in the linker region, and the assembly will interact with target antibodies via the Protein G binding region, to afford antibodies labelled with a long-lived fluorescence probe that can be visualised with time-gated techniques.