Despite
evidence showing that polymer brushes (PBs) are a powerful
tool used in biosensing for minimizing nonspecific interactions, allowing
for optimization of biosensing performance, and the fact that GaAs
semiconductors have proven to have a remarkable potential for sensitive
biomolecule detection, the combination of these two robust components
has never been considered nor evaluated as a platform for biosensing
applications. This work reports different methodologies to prepare
and tune PBs on the GaAs interface (PB–GaAs) and their potential
as useful platforms for antibody grafting, with the ultimate goal
of demonstrating the innovative and attractive character of the PB–GaAs
interfaces in the enhanced capture of antibodies and control of nonspecific
interactions. Three different functionalization approaches were explored,
one “grafting-to” and two “grafting-from,”
in which atom transfer radical polymerization (ATRP) was performed,
followed by their corresponding characterizations. Demonstration of
the compatibility of Escherichia coli (E. coli) and Legionella
pneumophila (Lp) antibodies with
the PB–GaAs platform compared to the results obtained with
conventional biosensing architectures developed for GaAs indicates
the attractive potential for operation of a sensitive biosensor. Furthermore,
these results showed that by carefully choosing the nature and preparation
methodology of a PB–GaAs interface, it is possible to effectively
tune the affinity of PB–GaAs-based sensors toward E. coli and Lp antibodies ultimately demonstrating
the superior specificity of the developed biosensing platform.
This work reports on the potential of polymer brushes
(PBs) grown
on GaAs substrates (PB-GaAs) as a promising platform for the detection
of
Legionella pneumophila
(Lp)
. Three functionalization approaches of the GaAs surface were used,
and their compatibility with antibodies against
Lp
was evaluated using Fourier transform infrared spectroscopy and
fluorescence microscopy. The incorporation of PBs on GaAs has allowed
a significant improvement of the antibody immobilization by increased
surface coverage. Bacterial capture experiments demonstrated the promising
potential for enhanced immobilization of
Lp
in comparison
with the conventional alkanethiol self-assembled monolayer-based biosensing
architectures. Consistent with an eightfold improved capture of bacteria
on the surface of a PB-functionalized GaAs/AlGaAs digital photocorrosion
biosensor, we report the attractive detection of
Lp
at 500 CFU/mL.
A regenerable bulk acoustic wave (BAW) biosensor is developed for the rapid, label-free and selective detection of Escherichia coli in liquid media. The geometry of the biosensor consists of a GaAs membrane coated with a thin film of piezoelectric ZnO on its top surface. A pair of electrodes deposited on the ZnO film allows the generation of BAWs by lateral field excitation. The back surface of the membrane is functionalized with alkanethiol self-assembled monolayers and antibodies against E. coli. The antibody immobilization was investigated as a function of the concentration of antibody suspensions, their pH and incubation time, designed to optimize the immunocapture of bacteria. The performance of the biosensor was evaluated by detection tests in different environments for bacterial suspensions ranging between 103 and 108 CFU/mL. A linear dependence between the frequency response and the logarithm of E. coli concentration was observed for suspensions ranging between 103 and 107 CFU/mL, with the limit of detection of the biosensor estimated at 103 CFU/mL. The 5-fold regeneration and excellent selectivity towards E. coli detected at 104 CFU/mL in a suspension tinted with Bacillus subtilis at 106 CFU/mL illustrate the biosensor potential for the attractive operation in complex biological media.
Since Legionella pneumophila has caused punctual epidemics through various water systems, the need for a biosensor for fast and accurate detection of pathogenic bacteria in industrial and environmental water has increased. In this report, we evaluated conditions for the capture of live L. pneumophila on a surface by polyclonal antibodies (pAb) and recombinant antibodies (recAb) targeting the bacterial lipopolysaccharide. Using immunoassay and PCR quantification, we demonstrated that, when exposed to live L. pneumophila in PBS or in a mixture containing other non-target bacteria, recAb captured one third fewer L. pneumophila than pAb, but with a 40% lower standard deviation, even when using the same batch of pAb. The presence of other bacteria did not interfere with capture nor increase background by either Ab. Increased reproducibility, as manifested by low standard deviation, is a characteristic that is coveted for biosensing. Hence, the recAb provided a better choice for immune adhesion in biosensors even though it was slightly less sensitive than pAb. Polyclonal or recombinant antibodies can specifically capture large targets such as whole bacteria, and this opens the door to multiple biosensor approaches where any of the components of the bacteria can then be measured for detection or characterisation.
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