Multiplexed point-of-care testing for inflammatory miRNA and neurotransmitter profiling in Alzheimer’s disease diagnostics

• Applicants: Professor Dr.-Ing. Can Dincer; Professor Dr. Michael Thomas Heneka
• Subject Area: Microsystems ; Biomedical Systems Technology
• Term: from 2019 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 421356369

• Final Report Year: 2024

Final Report Abstract

The scope of this project was the investigation of short (18-22 nucleotides) RNA sequences (miRNA) as potential biomarkers for immunological processes and neuroinflammation in Alzheimer’s disease (AD) alongside the implementation of a microfluidic device for their fast and sensitive detection. After pre-screening cerebrospinal fluid (CSF) samples obtained from a cohort of AD patients, as well as patients with other neurodegenerative diseases (University Hospital Bonn, UKB), multiple targets that had been reported as upregulated in literature, had to be dismissed, since they were not reproducibly quantifiable. Screening of a broader selection of miRNAs, we were able to identify miR-520f-3p as highest, immunologically relevant target in CSF. Hence, calibration of the biosensor for the detection of unamplified RNA (development and fabrication at the Institute for Microsystems Engineering, IMTEK, University of Freiburg) was conducted using synthetic miR-520f. Employing the optimized CRISPR-powered bioassay, we were able to achieve an analytical limit of detection of 3.6 fM in a sample-to-result time of approximately 30 min. The particularly high abundance of the target in RNA isolates derived from untreated THP-1 cells, that had been observed in the Nanostring screening could be reproduced and thus validated in amperometric measurements of the microfluidic sensor. In addition, we were able to design, implement and characterize a single-channel multiplexed version of the biosensor for simultaneous analysis and readout of up to six analytes.

Publications

• The impact of biosensing in a pandemic outbreak: COVID-19. Biosensors and Bioelectronics, 163, 112274.
  Morales-Narváez, Eden & Dincer, Can
  
• A CRISPR/Cas13a-powered catalytic electrochemical biosensor for successive and highly sensitive RNA diagnostics. Biosensors and Bioelectronics, 178, 113027.
  Sheng, Yan; Zhang, Tenghua; Zhang, Shihong; Johnston, Midori; Zheng, Xiaohe; Shan, Yuanyue; Liu, Tong; Huang, Zena; Qian, Feiyang; Xie, Zihui; Ai, Yiru; Zhong, Hankang; Kuang, Tairong; Dincer, Can; Urban, Gerald Anton & Hu, Jiaming
  
• CRISPR-powered electrochemical microfluidic multiplexed biosensor for target amplification-free miRNA diagnostics. Biosensors and Bioelectronics, 177, 112887.
  Bruch, Richard; Johnston, Midori; Kling, André; Mattmüller, Thorsten; Baaske, Julia; Partel, Stefan; Madlener, Sibylle; Weber, Wilfried; Urban, Gerald A. & Dincer, Can
  
• Current Technologies for RNA-Directed Liquid Diagnostics. Cancers, 13(20), 5060.
  Martinez-Dominguez, Maria Victoria; Zottel, Alja; Šamec, Neja; Jovčevska, Ivana; Dincer, Can; Kahlert, Ulf Dietrich & Nickel, Ann-Christin
  
• Disposable Intergrated Sensors for Next-generation On-site Testing, AsiaSense, 21.10.2021
  Johnston M., Ates H.C., Laubender E., Bruch R., Maier D., Baaske J., Weber W., Urban GA & Dincer C.
  
• Integrated sensing devices for point-of-care diagnostics, Sensors in Medicine, 9.11.2021
  Ates H.C., Johnston M., Laubender E., Bruch R., Maier D., Baaske J., Weber W., Urban G.A. & Dincer C.
  
• Non‐Invasive Diagnostics: Integrated Devices for Non‐Invasive Diagnostics (Adv. Funct. Mater. 15/2021). Advanced Functional Materials, 31(15).
  Ates, Hatice Ceren; Brunauer, Anna; von Stetten, Felix; Urban, Gerald A.; Güder, Firat; Merkoçi, Arben; Früh, Susanna Maria & Dincer, Can
  
• CRISPR/Cas-powered nanobiosensors for diagnostics. Biosensors and Bioelectronics, 197, 113732.
  Phan, Quynh Anh; Truong, Linh B.; Medina-Cruz, David; Dincer, Can & Mostafavi, Ebrahim
  
• Designing electrochemical microfluidic multiplexed biosensors for on-site applications. Analytical and Bioanalytical Chemistry, 414(22), 6531-6540.
  Glatz, Regina T.; Ates, H. Ceren; Mohsenin, Hasti; Weber, Wilfried & Dincer, Can
  
• Multiplexed biosensor for point-of-care COVID-19 monitoring: CRISPR-powered unamplified RNA diagnostics and protein-based therapeutic drug management. Materials Today, 61, 129-138.
  Johnston, Midori; Ceren, Ates H.; Glatz, Regina T.; Mohsenin, Hasti; Schmachtenberg, Rosanne; Göppert, Nathalie; Huzly, Daniela; Urban, Gerald A.; Weber, Wilfried & Dincer, Can
  
• Electrochemical microfluidic multiplexed biosensor towards CRISPR-powered detection of unamplified RNA and protein-based therapeutic drug management, 33rd Anniversary World Congress on Biosensors, 8. Juni, 2023, Busan, Korea (Präsentation)
  Johnston M., Ates H.C., Glatz R.T., Mohsenin H., Schmachtenberg R., Göppert N., Huzly D., Urban G., Weber W. & Dincer C.