Development of Low-cost and High Sensitive Platform for Rapid Disease Diagnosis

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In order to manage and treat diseases rapidly, it is important to identify the cause of a disease accurately and quickly. In particular, emerging infectious diseases and cancers are the main causes of death, and treatment and management differ depending on the presence of targeted pathogens or mutations. For these reasons, accurate diagnosis is necessary. Currently developed diagnostic methods are high-cost, time-consuming methods with low sensitivity and specificity. In addition, in cases of emerging infectious diseases, the current method of diagnosis requires complex equipment and technology, despite the need for rapid on-site diagnosis. To overcome these limitations, rapid and accurate disease diagnostic approach was developed using a sample preparation technology-based microfluidic platform and bio-optical sensor based silicon microring resonator.
For clinical sample preparation, a microfluidic platform was developed that enabled pathogen enrichment and nucleic acids extraction using homobifunctional imidoester reagents. HI reagents bind to pathogens or nucleic acids via covalent bonding and electrostatic interactions. To validate the clinical utility of this platform and compare its performance with that of conventional methods, we obtained plasma samples from patient with ST and SFTS patients, 10 nasopharyngeal samples from patient with HAdV infection, 12 environmental swab samples exposed to patients with hPIV-3, and 20 saliva and 14 plasma samples from paateints with HZ. A microfluidic platform that can isolate cell-free DNA from the plasma of patients with cancer without a cell lysis step was also developed. cfDNA isolated using this platform can be analyzed oncogenic mutations (BRAF, KRAS). Moreover, the Oncopanel result of isolated cfDNA compared to WES result of tissue specimens from patient with colorectal cancer. Finally, for detecting nucleic acids, a bio-optical sensor using the isothermal PCR method was developed. This sensor was validated using DNA from patients with ADV, HZ and genomic DNA or cfDNA from patients with cancer.
Sample preparation using a microfluidic platform could handle various clinical samples within 50 min using one chip. This platform can increase the enrichment ratio more than 80% compared to that with a conventional kit. Its clinical utility in large sample volumes was demonstrated in 46 clinical specimens including environmental swabs, saliva, and blood plasma. This system showed higher sensitivity with these samples and could detect pathogens that were below the threshold of detection using other methods. Moreover, after isolation of cfDNA from the plasma of patients with cancer using the microfluidic platform, the detection efficiency of circulating tumor DNA was increased because of decreased background DNA compared to that using a conventional kit. When the WES result of tissue from patients was compared, the concordance ratio of mutations was 71.4%. With the increasing sensitivity of ctDNA detection, oncogenic mutations in cfDNA can be identified using sequencing or PCR without high-cost NGS analysis. Last, when detecting pathogens or mutations using the bio-optical sensor, the sensitivity of detection is 10-100 times higher compare to that of the qPCR method. By combining the sample preparation using the microfluidic platform, the possibility of a fast and highly sensitive diagnostic platform was confirmed.
In this study, a microfluidic platform enabling sample preparation within 50 min was integrated with a bio-optical sensor for detecting nucleic acids within 20 min. This integrated assay led to rapid and sensitive disease diagnosis technology in clinical specimens. Integrated systems such as this have considerable potential as POC based pathogen diagnostic systems that could have diverse clinical applications in humans and also in animal healthcare. It could also be useful for the clinical diagnosis and monitoring of cancer and infectious disease treatment.
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Jin, Choong Eun
일반대학원 의학과의공학전공
Shin, Yong
울산대학교 일반대학원 의학과의공학전공
울산대학교 논문은 저작권에 의해 보호받습니다.
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Medical Engineering > 2. Theses (Ph.D)
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