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Identifier

000417585

Title

Development of acoustic wave biosensors for application in molecular diagnostics

Alternative Title

Ανάπτυξη ακουστικών βιοαισθητήρων με εφαρμογή στη μοριακή διαγνωστική

Author

Γραμμουστιανού, Αριστέα Δ.

Thesis advisor

Γκιζελή, Ηλέκτρα

Reviewer

Βόντας, Ιωάννης
Καλαντίδης, Κρίτων

Abstract

Since the development of the first biosensor by Leland C. Clark in 1962, the progress in the field of biosensors has been phenomenal. The biosensors market is estimated to reach US$ 27.06 billion by the year 2022 and, currently, scientific research is focused on the development of inexpensive, non-invasive, integrated and portable biosensors that will find application in point-of-care diagnostics, food and process control, environmental monitoring, biodefense, and forensics. The present PhD research focused on the development of acoustic wave-based bioassays that can be applied in molecular diagnostics. Acoustic wave biosensors offer real-time and label-free biological sensing. This study made use of the unique characteristics of acoustic wave biosensors in order to develop bioassays for food safety and cancer diagnostics. The first objective of this thesis was the acoustic detection of the foodborne pathogen Salmonella Typhimurium. Two approaches were followed: the first one made use of an acoustic wave immunosensor for the detection of whole Salmonella Typhimurium cells. The biosensor developed was able to detect Salmonella Typhimurium cells of a concentration of 10⁶ cfu/mL and within 30 min. The second approach employed an acoustic wave biosensor for the solid phase amplification of the gene invA of Salmonella Typhimurium via the recombinase polymerase amplification and simultaneous detection of DNA amplicons on the device surface. For amplicon detection, a novel methodology was described, where the formation of surface-bound DNA amplicons was detected by using liposomes as an amplification probe. The presented methodology was fast (detection of DNA amplicons after only 5 min of amplification) and simple (it did not require any post-amplification steps). The second part of the research work focused on the development of acoustic wave biosensors for molecular cancer diagnostics. A DNA-based biosensor was designed for the detection of four miRNAs (miR-21, miR-155, miR-150 and miR-107) considered being dynamic biomarkers of breast cancer. The miRNAs detection was achieved using a single oligonucleotide probe. In fact, the biosensor allowed the rapid, real-time, label-free and simple detection of two miRNAs at the same time. The final aim of this work was the development of an acoustic wave-based bioassay by which ctDNA carrying KRAS mutations that are characteristic of colorectal cancer would be acoustically detected via their hybridization to a PNA probe immobilized on the acoustic sensor surface; the assay employed a high fundamental frequency acoustic wave device and a patented technology which combines the patented aldehyde-modified SMART nucleobases with unique PNA capture probes synthesized with a “blank” position (DGL probes). This technology introduces an entirely chemical, rather than enzymatic, method for nucleic acid testing. The assay achieved the detection of KRAS mutations present on ctDNA introducing a new alternative to current colorectal cancer diagnostic techniques.

Physical description

Ορκομωσία??

Language

English

Subject

Biosensing

Cancer diagnostics

Food safety

QCM

SAN

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