IMPEDANCE BASED APTASENSOR FOR THE DETECTION OF MYCOBACTERIUM TUBERCULOSIS SECRETED PROTEIN MPT64
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Date
2019
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Nazarbayev University School of Engineering and Digital Sciences
Abstract
Tuberculosis (TB) detection remains a significant healthcare issue in the developing world
owing to a number of challenges. Current diagnostics are based on microbiological culturing,
sputum smear microscopy, and nucleic acid amplification tests. These methods suffer from
limitations such as batch to batch variations, frequent contaminations, low sensitivity, and the
requirement for special facilities, expensive devises, reagents, and trained personnel.
This thesis describes the development of the sensitive oligonucleotide-based aptasensor for the
detection of TB biomarker MPT64 protein. The dissertation investigates the selection and use
of ssDNA aptamers to detect MPT64 using the electrochemical impedance spectroscopy (EIS).
Aptamers serve as bio-recognition elements in this study, and they have numerous advantages
including cheap cost, ease of modification and long shelf life. The combination of aptamers
with the EIS offers sensitive detection since the change in EIS signal can be recorded as the
result of analyte binding event based not only on molecular interaction level but also on
electron transfer levels.
As the result 17 unique aptamer sequences were purified and analyzed. One aptamer with
dissociation equilibrium constant KD of 8.92 nM was selected and the surface chemistry was
optimized based on ssDNA aptamer modified with a long linker and 6-mercaptohexanol as a
co-adsorbent at 1/100 ratio. The selected aptamer was further immobilized on an interdigitated
microelectrode and connected to a portable potentiostat. The detection time for aptasensor was
found to be 15 min. The aptasensor was tested on clinical samples and showed increased
binding to TB (+) samples as compared to TB (-) samples. The integration of the aptasensor
with the in house built fluidic chamber and liquid flow rate within chamber was also
investigated.
The work in this thesis is significant as it can contribute to the diagnosis of TB (non-invasive),
monitoring of anti-TB treatment in infected people and hence to socio-economic development
of the country. It is the first portable aptasensor which is developed using aptamers and EIS as
a detection technique that can provide fast clinical sample analysis (reduced from 3 h to 15
min) as well as elimination of using of extra reagents, equipment, and personnel.
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Citation
Sypabekova, M. (2019). Impedance Based Aptasensor For The Detection Of Mycobacterium Tuberculosis Secreted Protein Mpt64. Nazarbayev University School of Engineering and Digital Sciences