Quantum dots (QDs) are semiconductor nanoparticles which have emerged as powerful
fluorescent probes for biological imaging applications due to their unique size-dependent
optical and electrical properties. QDs have several advantages over small organic dyes and
fluorescent proteins such as size-tunable photoluminescence, wide excitation-narrow
emission properties, improved brightness and high resistance to photobleaching and
degradation. So far QDs have been used to track individual biomolecules, but for this
application a widespread concern is that biomolecules can lose activity when they are
attached to QDs because these are multivalent and large. Thus, recent attention has turned
toward labelling strategies which enable site-specific recognition and controlling the number
of molecules that can be attached to a single QD down to a single molecule with retention of
activity. Apart from showing ability to recognise appropriate biological partners, relatively
little is known about the biological activity of biomolecules attached to QDs.
In this thesis various strategies for preserving and enhancing the activity of
biomolecules on QDs were developed to address and investigate these aspects and to extend
the biological applications of QDs.
Nitrilotriacetic acid (NTA)-modified QDs were used for site specific labelling o f a
hexahistidine (His₆)-tagged Glutathione-S-Transferase (GST). GSTs catalyse nucleophilic
substitution reactions between glutathione and a wide range of endogenous and xenobiotic
electrophiles, which makes them important detoxifying enzymes and anticancer targets. The
hydrophobic CdSe-ZnS (core-shell) QDs were made water soluble by ligand exchange with
dihydrolipoic acid and coupled to NTA-Ni via an amide bond. Ni-NTA capped QD were
capable of binding recombinant S. japonicum His6-GST selectively. As a result of the His₆
tag’s ability to provide a docking site for the QD away from the active site, the GST
molecules bound to these QDs retained their catalytic activity. In contrast, the non specific
binding which takes place in the absence of the His₆ tag leads to loss o f catalytic activity.
Hydrophobic interactions were used to functionalize CdSe-ZnS QDs with Kdo2-lipid A
-the lipopolysaccahride (LPS) present in the outer membrane of E.coli. These constructs
were used as pathogen models to investigate how pathogens and pathogen associated
molecular patterns (e.g. LPS) interact and are processed by the immune system. The ability of
QDs to enhance the biological activity o f a biomolecule was demonstrated in vitro and in vivo
for the first time. QD-LPS micelles were able to induce stronger production of cytokines in
macrophages and dendritic cells in vitro and a model antigen (DNP-OVA) in vivo than
Also presented in this thesis is the first attempt to exploit the multivalency and site
specific labelling properties of NTA-Ni-decorated QDs to mimic the surface o f a parasite.
The focus here was on the Plasmodium falciparum malaria merozoite, which has MSP 1 as
major component o f its surface. Conjugation of a recombinant form o f His6-MSP-l hybrid to
three different types o f NTA-Ni-decorated QDs was accomplished. Morever, by changing the
linker units separating the QDs and Ni-NTA complexes it was possible to control the number
of MSP 1 molecules attached to each QD.