Development of a controller for gene‐expression analysis using field-effect devices

• Autor(a) principal: Sá, André Gomes de
• Data de Publicação: 2019
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10362/90040
• topic: Transimpedance amplifier; analog-front-end circuit; LAMP; pH; Domínio/Área Científica::Engenharia e Tecnologia::Nanotecnologia

Field-effect-based devices have become a basic structural element in a new generation of biosensors and are today the foundation of radical new approaches for specific detection and characterization of DNA. Reliable molecular detection and characterization of genetic biomarkers are vital for disease diagnostics and therapeutic follow-up. In this work, an analog front-end circuit (AFE) was developed for an ion-sensitive FET- based real-time monitorization of isothermal DNA amplification. For this, a microcontroller/circuit integration was developed allowing for precise control of applied voltage, enabling real-time output measurements. This prototype was built over an Arduino microcontroller and was developed as a standalone platform with dedicated software for control and data handling, for a sample in result out- like sensing platform. This prototype as shown to be able to detect changes in charge/pH comparable to previously attained results measured in a standard apparatus (22± 6 mV/pH – 58mv/pH theoretical maximum via Nernst Equation). The platform was tested with standard pH solutions (ranging from 4 to 12) and was able to convert, amplify and monitor the current variation in the order of 0.3μA/pH generated by the surface potential difference. This approach was integrated with isothermal amplification techniques (LAMP) for a relevant cancer biomarker (c-Myc proto-oncogene). Real-time LAMP amplification reactions were monitored using the standard fluorescence-based approach and compared to those attained via the developed sensing platform. Results show that the proposed platform allows for the real-time monitorization of LAMP amplification. At this stage, the error associated with the measurement increases over time, with decreasing signal to noise ratios. Also, a noticeable change in reaction kinetics was observed, leading to an increase in reaction times. Subsequently, the proposed circuit was implemented in a printed circuit board in order to reduce noise and improve overall stability.