Jon Zabalo Carrere
Diseño, fabricación, caracterización y validación de una plataforma integrada y modular para la amplificación de secuencias específicas de ADN por PCR basada en dispositivos de microfluídica digital.
Degree: Biomedical Engineer
Thesis Director: Sergio Arana Alonso
The Polymerase Chain Reaction (PCR) is one of the most well-known molecular biology techniques used in the diagnosis of diseases and detection of organisms in different types of samples. This technique allows for the creation of multiple copies of a specific DNA sequence by performing several cycles composed of stages at different temperatures. This way, starting from a small amount of sample to be analyzed, enough replicas of the DNA fragment of interest are created to make it analyzable or detectable in detail. Among the main applications of PCR are the detection of infectious diseases such as HIV or SARS-CoV-2, the detection of DNA mutations that may cause diseases such as cancer, and the detection of pathogens or traces of allergens in different foods.
PCR reactions are usually carried out in laboratory equipment known as thermal cyclers, which are composed of a heating element that thermally cycles the samples to be analyzed. However, these instruments require temperature ramps and relatively large sample volumes to perform the reactions. In response to this problem, the emergence of lab-on-a-chip technology and microfluidics have accelerated the development of miniaturized systems to carry out various molecular biology analyses more efficiently.
In particular, digital microfluidics (DMF) offers the advantages of conventional microfluidic systems such as shorter reaction times, lower reagent consumption, reduced sample contamination, and improved sensitivity and overall efficiency of the system. In addition, DMF has additional advantages over conventional microfluidics, including the absence of microchannels and liquid propulsion pumps, high transfer speeds of samples, rapid heat transfer, and the absence of void or "dead" volumes in the devices. This is because DMF technology allows for the automated control of discrete sample volumes along an array of actuator electrodes thanks to electrostatic forces.
With the aim of creating an efficient PCR system, this work has developed a modular digital microfluidic platform that allows for the integration of different DMF devices for nucleic acid amplification by PCR. For this purpose, DMF devices were designed and fabricated using microfabrication techniques, and after integrating them into the developed DMF platform, various studies were carried out. Firstly, DMF devices were developed for studies of the dynamic behavior of the proposed microfluidic system. Then, DMF devices focused on performing PCR reactions, with integrated resistive heaters, were developed, which made it possible to carry out nucleic acid amplification by PCR more efficiently and satisfactorily compared to conventional PCR systems, also validating the developed DMF system with real samples.