Microfluidic devices with integrated biosensors for biomedical applications

Abstract

In recent years, the LOC community has focused most of its research in the biomedical and biotechnology fields, due to the need of portable, low power consumption and low cost theranostics microdevices. Some developing countries do not have suitable medical diagnostics technologies and the supply and storage of the reagents is in many cases limited as well as the access to energy. Furthermore, developed countries are experimenting population aging needing novel low cost efficient disease-screening technologies. The introduction of LOC and microfluidics allow the integration of complex functions that could lead to the developing of more accurate, cheap and reliable theranostic tools. Current focus of application is focused mostly in drug delivery 1, cellular analysis 2, and disease diagnosis 3. Microfluidics is improving the developing of novel point-of-care devices, but there are some challenges that are slowing down the massive production of these LOC. These areas include new methods for sample collection, world-to-chip interfaces, sample pre-treatment, improvement of long-term stability of reagents, working with complex sample specimens, multiple detection of biomarkers and simplify the read-out 4. The main aim of this thesis work was to create novel, cheap and with a high degree of automatization miniaturized biosensing devices with the objective to facilitate Point-of-Care diagnostics in the near future. Our efforts have been focused into developing a LOC system with electrochemical sensing capabilities adjustable to any biomarker, depending only on sample volumes and required analysis times. The devices integrate low-cost label-free biosensors exploiting microfluidics-based self-functionalization, or specialization. The biosensor functionalization takes place in situ and selectively, just before the sensing, and their area keeps dry and inactive until the test starts. The reagents and the sensing parts are kept separated and brought into contact just before the test, avoiding the need of complex fabrication and storage methods to guarantee functionalization integrity. The novel design reduces the cost of the final instrumentation, by simplifying the measurements, while keeping sensitivities and LODs relevant for the application. Furthermore, since the interaction of antibody and protein is time and concentration dependent, our device has the capability to adjust its sensitivity. We have tuned and characterized our system sensitivity using different biomarkers. The development of our novel devices was possible by exploiting synergies in disciplines previously studied in our group. Particularly, in fields such as microfluidics 5-8, surface functionalization 9-14 and electrochemical biosensors 15-19. Summarizing, we are proposing novel microfluidic devices with integrated biosensors. The systems are based on the principle of laminar co-flow in order to perform an on-chip selective surface bio-functionalization of LOC integrated biosensors. This method has the advantage of performing the surface modification protocols “in situ” before the detection. The system can be easily scaled to incorporate several sensors with different biosensing targets in a single chip. We are proposing a novel voltage and impedance differential measurements; that allow us to simplify the read-out. As biomedical application we focus our attention on the detection of prostate cancer biomarkers. Bibliography 1. I. U. Khan, C. A. Serra, N. Anton and T. Vandamme, Journal of Controlled Release, 2013, 172, 1065-1074. 2. H. Andersson and A. Van den Berg, Sensors and Actuators B: Chemical, 2003, 92, 315-325. 3. M. J. Cima, Annual Review of Chemical and Biomolecular Engineering, 2011, 2, 355-378. 4. C. D. Chin, V. Linder and S. K. Sia, Lab on a Chip, 2012, 12, 2118-2134. 5. R. Rodriguez-Trujillo, C. A. Mills, J. Samitier and G. Gomila, Microfluidics and Nanofluidics, 2007, 3, 171-176. 6. R. Rodriguez-Trujillo, O. Castillo-Fernandez, M. Garrido, M. Arundell, A. Valencia and G. Gomila, Biosensors and Bioelectronics, 2008, 24, 290-296. 7. O. Castillo-Fernandez, R. Rodriguez-Trujillo, G. Gomila and J. Samitier, Microfluidics and Nanofluidics, 2014, 16, 91-99. 8. J. Comelles, V. Hortigüela, J. Samitier and E. Martínez, Langmuir, 2012, 28, 13688-13697. 9. E. Prats-Alfonso, F. García-Martín, N. Bayo, L. J. Cruz, M. Pla-Roca, J. Samitier, A. Errachid and F. Albericio, Tetrahedron, 2006, 62, 6876-6881. 10. J. Vidic, M. Pla-Roca, J. Grosclaude, M.-A. Persuy, R. Monnerie, D. Caballero, A. Errachid, Y. Hou, N. Jaffrezic-Renault, R. Salesse, E. Pajot-Augy and J. Samitier, Analytical Chemistry, 2007, 79, 3280-3290. 11. Y. Hou, S. Helali, A. Zhang, N. Jaffrezic-Renault, C. Martelet, J. Minic, T. Gorojankina, M.-A. Persuy, E. Pajot-Augy, R. Salesse, F. Bessueille, J. Samitier, A. Errachid, V. Akimov, L. Reggiani, C. Pennetta and E. Alfinito, Biosensors and Bioelectronics, 2006, 21, 1393-1402. 12. S. Rodríguez Seguí, M. Pla, J. Minic, E. Pajot‐Augy, R. Salesse, Y. Hou, N. Jaffrezic‐Renault, C. A. Mills, J. Samitier and A. Errachid, Analytical Letters, 2006, 39, 1735-1745. 13. A. Lagunas, J. Comelles, E. Martínez and J. Samitier, Langmuir, 2010, 26, 14154-14161. 14. A. Lagunas, J. Comelles, S. Oberhansl, V. Hortigüela, E. Martínez and J. Samitier, Nanomedicine: Nanotechnology, Biology and Medicine, 2013, 9, 694-701. 15. M. Castellarnau, N. Zine, J. Bausells, C. Madrid, A. Juárez, J. Samitier and A. Errachid, Materials Science and Engineering: C, 2008, 28, 680-685. 16. M. Castellarnau, N. Zine, J. Bausells, C. Madrid, A. Juárez, J. Samitier and A. Errachid, Sensors and Actuators B: Chemical, 2007, 120, 615-620. 17. M. Kuphal, C. A. Mills, H. Korri-Youssoufi and J. Samitier, Sensors and Actuators B: Chemical, 2012, 161, 279-284. 18. D. Caballero, E. Martinez, J. Bausells, A. Errachid and J. Samitier, Analytica Chimica Acta, 2012, 720, 43-48. 19. M. Barreiros dos Santos, J. P. Agusil, B. Prieto-Simón, C. Sporer, V. Teixeira and J. Samitier, Biosensors and Bioelectronics, 2013, 45, 174-180.

En años recientes, la comunidad de LOC ha enfocado todos sus esfuerzos en la investigación de nuevas aplicaciones para la biomedicina y biotecnología. Algunos países en vías de desarrollados no tienen tecnologías de diagnóstico adecuadas, además el suministro y almacenamiento de los reactivos es en muchos casos limitado, y en ocasiones cuentan con un acceso limitado al consumo de energía. Por otra parte, los países desarrollados se han encontrado con una población envejecida, y por lo tanto se ha generado la necesidad de contar con nuevas tecnologías para el diagnóstico de enfermedades las cuales sean accesibles y orientadas a una terapia más personalizada. Tanto la microfluídica como los LOC han permitido la integración de funciones de análisis complejas capaces de desarrollar herramientas de diagnostico más precisas, de bajo coste y confiables. Actualmente toda la atención se ha centrado en el diseño de aplicaciones para administración de fármacos 1, análisis celular 2 y diagnostico de enfermedades 3. La introducción de la microfluídica ha servido para mejorar el desarrollo de nuevos dispositivos point-of-care, pero todavía existen algunos problemas que han evitado la producción masiva de estos LOC. Las áreas en las que se pretende conseguir una mejora son la recolección de la muestra, mejora de la interfaz entre el chip y el usuario, tratamiento previo de la muestra, mejorar la estabilidad de los reactivos, trabajo con muestras complejas, detección múltiple de biomarcadores y simplificación del sistema de medida 4. Nuestros esfuerzos se han dedicado en desarrollar un sistema LOC con capacidad de detección electroquímica ajustable a cualquier biomarcador, dependiendo únicamente en la cantidad de muestra y los tiempos de análisis. Nuestros dispositivos microfluídicos cuentan con biosensores integrados de bajo coste con capacidad de auto-funcionalización. La funcionalización de los biosensores se realiza in-situ y selectivamente, antes de la detección, manteniendo el área de detección inerte hasta el inicio de la prueba. Los reactivos y el área de detección se almacenan por separado y entran en contacto hasta el inicio del experimento, lo cual facilita el método de fabricación. Se ha podido desarrollar este trabajo gracias a los estudios previos realizados en nuestro grupo en distintas disciplinas, tales como: microfluídica 5-8, funcionalización de superficies 9-14 y biosensores electroquímicos 15-19. Bibliografía 1. I. U. Khan, C. A. Serra, N. Anton and T. Vandamme, Journal of Controlled Release, 2013, 172, 1065-1074. 2. H. Andersson and A. Van den Berg, Sensors and Actuators B: Chemical, 2003, 92, 315-325. 3. M. J. Cima, Annual Review of Chemical and Biomolecular Engineering, 2011, 2, 355-378. 4. C. D. Chin, V. Linder and S. K. Sia, Lab on a Chip, 2012, 12, 2118-2134. 5. R. Rodriguez-Trujillo, C. A. Mills, J. Samitier and G. Gomila, Microfluidics and Nanofluidics, 2007, 3, 171-176. 6. R. Rodriguez-Trujillo, O. Castillo-Fernandez, M. Garrido, M. Arundell, A. Valencia and G. Gomila, Biosensors and Bioelectronics, 2008, 24, 290-296. 7. O. Castillo-Fernandez, R. Rodriguez-Trujillo, G. Gomila and J. Samitier, Microfluidics and Nanofluidics, 2014, 16, 91-99. 8. J. Comelles, V. Hortigüela, J. Samitier and E. Martínez, Langmuir, 2012, 28, 13688-13697. 9. E. Prats-Alfonso, F. García-Martín, N. Bayo, L. J. Cruz, M. Pla-Roca, J. Samitier, A. Errachid and F. Albericio, Tetrahedron, 2006, 62, 6876-6881. 10. J. Vidic, M. Pla-Roca, J. Grosclaude, M.-A. Persuy, R. Monnerie, D. Caballero, A. Errachid, Y. Hou, N. Jaffrezic-Renault, R. Salesse, E. Pajot-Augy and J. Samitier, Analytical Chemistry, 2007, 79, 3280-3290. 11. Y. Hou, S. Helali, A. Zhang, N. Jaffrezic-Renault, C. Martelet, J. Minic, T. Gorojankina, M.-A. Persuy, E. Pajot-Augy, R. Salesse, F. Bessueille, J. Samitier, A. Errachid, V. Akimov, L. Reggiani, C. Pennetta and E. Alfinito, Biosensors and Bioelectronics, 2006, 21, 1393-1402. 12. S. Rodríguez Seguí, M. Pla, J. Minic, E. Pajot‐Augy, R. Salesse, Y. Hou, N. Jaffrezic‐Renault, C. A. Mills, J. Samitier and A. Errachid, Analytical Letters, 2006, 39, 1735-1745. 13. A. Lagunas, J. Comelles, E. Martínez and J. Samitier, Langmuir, 2010, 26, 14154-14161. 14. A. Lagunas, J. Comelles, S. Oberhansl, V. Hortigüela, E. Martínez and J. Samitier, Nanomedicine: Nanotechnology, Biology and Medicine, 2013, 9, 694-701. 15. M. Castellarnau, N. Zine, J. Bausells, C. Madrid, A. Juárez, J. Samitier and A. Errachid, Materials Science and Engineering: C, 2008, 28, 680-685. 16. M. Castellarnau, N. Zine, J. Bausells, C. Madrid, A. Juárez, J. Samitier and A. Errachid, Sensors and Actuators B: Chemical, 2007, 120, 615-620. 17. M. Kuphal, C. A. Mills, H. Korri-Youssoufi and J. Samitier, Sensors and Actuators B: Chemical, 2012, 161, 279-284. 18. D. Caballero, E. Martinez, J. Bausells, A. Errachid and J. Samitier, Analytica Chimica Acta, 2012, 720, 43-48. 19. M. Barreiros dos Santos, J. P. Agusil, B. Prieto-Simón, C. Sporer, V. Teixeira and J. Samitier, Biosensors and Bioelectronics, 2013, 45, 174-180.