Development of impedimetric immunosensor based on modified pt disk microelectrode for detection of amyloid beta in brain tissue lysate / Norazreen Zakaria
Alzheimer's disease (AD) is a fatal neurodegenerative disease and clinically characterized by progressive deterioration of memory and decline of cognitive function. Since the only reliable diagnosis for the pathology is histochemical post-mortem examination, therefore, there is a continuing dem...
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| Format: | Thesis |
| Language: | English |
| Published: |
2018
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| Online Access: | https://ir.uitm.edu.my/id/eprint/82540/1/82540.pdf |
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| Summary: | Alzheimer's disease (AD) is a fatal neurodegenerative disease and clinically characterized by progressive deterioration of memory and decline of cognitive function. Since the only reliable diagnosis for the pathology is histochemical post-mortem examination, therefore, there is a continuing demand for fast, reliable, and sensitive detection technique. Here, a miniaturized biosensing platform based on monoclonal amyloid-beta antibodies (mAβab) that were immobilized on a disc-shaped platinum/iridium (Pt/Ir) microelectrode surface coupled with an impedimetric signal transducer has been developed for a label-free and sensitive detection of amyloid-beta peptide fragment 1−40 (Aβ40), a reliable biomarker for diagnosis of AD. The Pt/Ir microelectrode was electropolymerized with poly (ortho-phenylenediamine), a conducting free amine-containing aromatic polymer; followed by crosslinking with glutaraldehyde (GA) for subsequent coupling of mAβab on the microelectrode surface. This modification strategy efficiently improved the impedimetric detection performance of Aβ40 in terms of charge transfer resistance and normalized impedance magnitude percentage change (~40 % increase) as compared to passive adsorption-based immobilization method. The sensitivity of the micro-immunosensing assay was 1056 pg/mL/ (kΩ•cm2) and the limit of detection of 4.81 pg/mL with a dynamic range of 1−105 pg/mL (R2=0.9932) were obtained. The repeatability of the assay via relative standard deviation was 12.8 %, demonstrating its reliability and accuracy. In respect of sensor durability and stability, the immobilized mAβab on the microelectrode surface was capable of maintaining 80 % of its binding activity to Aβ40 after leaving in PBS (pH 7.4) solution for 48 h at ambient temperature. To validate the real applicability, the developed assay was tested in brain tissue lysates prepared from AD-induced rats. The results were presented which includes the quantification data of Aβ40 presence in the samples. As a conclusion, this study demonstrates the potential application of the Pt-PPD-GA-mAβab-Aβ40 immunosensing assay for diagnosing the AD‘s biomarker. The method used in this work would offer a useful means for quantifying Aβ in a biological matrix, and be valuable in the design of new types of electrochemical biosensors for the detection of other disease-related biomarkers. |
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