Periodic Reporting for period 2 - Smart BioSense (Smart engineered Bioreceptors for developing BioSensors)
Okres sprawozdawczy: 2020-11-01 do 2021-10-31
Although the impressive advances in the field of biosensing, most efforts are still oriented to deliver lower limits of detection by updating or improving either the molecular reporters or the reading set ups, while little effort is addressed towards the recognition element/bio receptor. Better insight in the recognition element biophysics allow to tune its properties to match the needs for the receptor in each particular context. These improvements in receptors’ properties have a direct impact and benefit on any biosensing assay technology assay, not only the new developed, but also in previously established techniques.
The action was divided into three main objectives:
1. Exploration and optimization of binding conformational changes as a signal transduction mechanism.
2. Rational introduction of cooperativity to improve measurement precision.
3. Demonstration of the application of these mechanism in representative biosensors.
In the MSCA action, the experienced researcher (Dr. Alejandro Chamorro Garcia) spent the outgoing phase in the host institution (University of California Santa Barbara, UCSB) in Prof. Kevin Plaxco research lab. The work carried out during outgoing phase the work had been centered in the use of diverse mechanisms, such as cooperativity and sequestration, to tune biological receptors’ recognition properties towards the application of the improved receptors in biosensing assays. With the final goal to achieve improved response from the sensing assays in which are applied, we focused on narrowing the dynamic range to obtain more precise reads of subtle changes in concentrations, in an on/off fashion. In the incoming phase the experience researched joined Prof. Francesco Ricci lab (University of Rome Tor Vergata) to further pursue the study of mechanisms to tune binding properties of bioreceptors and its applications in the development of biosensors.
In conclusion, during the action, the experience researcher had received training in electrochemical aptamer-based biosensors (EAB sensors) and biophysics of biological receptors. The knowledge acquired has been put into practice by studying the effect of the position of the label in the aptamer in EAB sensors and developing an EAB sensor for the detection of a chemotherapeutical drug: methotrexate. In another line, the experienced researcher applyed a previously described principle (sequestration) to tune receptors’ binding properties with the aim to improve the dose response parameters. That means inducing bigger transitions of signal over target concentration, achieving this way, better sensitivity over subtle variations in concentration. The principles developed and studies have been tested in few example testing scenarios, with the potential to be exploited in other contexts and help other researchers to incorporate improvements in their sensing systems.
The action was divided into three main objectives:
1. Exploration and optimization of binding conformational changes as a signal transduction mechanism.
2. Rational introduction of cooperativity to improve measurement precision.
3. Demonstration of the application of these mechanism in representative biosensors.
In the MSCA action, the experienced researcher (Dr. Alejandro Chamorro Garcia) spent the outgoing phase in the host institution (University of California Santa Barbara, UCSB) in Prof. Kevin Plaxco research lab. The work carried out during outgoing phase the work had been centered in the use of diverse mechanisms, such as cooperativity and sequestration, to tune biological receptors’ recognition properties towards the application of the improved receptors in biosensing assays. With the final goal to achieve improved response from the sensing assays in which are applied, we focused on narrowing the dynamic range to obtain more precise reads of subtle changes in concentrations, in an on/off fashion. In the incoming phase the experience researched joined Prof. Francesco Ricci lab (University of Rome Tor Vergata) to further pursue the study of mechanisms to tune binding properties of bioreceptors and its applications in the development of biosensors.
In conclusion, during the action, the experience researcher had received training in electrochemical aptamer-based biosensors (EAB sensors) and biophysics of biological receptors. The knowledge acquired has been put into practice by studying the effect of the position of the label in the aptamer in EAB sensors and developing an EAB sensor for the detection of a chemotherapeutical drug: methotrexate. In another line, the experienced researcher applyed a previously described principle (sequestration) to tune receptors’ binding properties with the aim to improve the dose response parameters. That means inducing bigger transitions of signal over target concentration, achieving this way, better sensitivity over subtle variations in concentration. The principles developed and studies have been tested in few example testing scenarios, with the potential to be exploited in other contexts and help other researchers to incorporate improvements in their sensing systems.
In the first stage of the action, the host institution provided the experienced researcher with a strong background in EAB sensors’ fabrication and use. The work and training during this package lead to the study of alternate labelling of aptamer used in the EAB sensors. We explored the alternative placing of the functionalities in the aptamers for EDNA sensors achieving results that were published in a scientific journal. In addition, the knowledge and experience acquired was ultimately applied by developing and characterizing a new EAB sensor, not reported before, for the detection of a chemotherapeutic drug (methotrexate).
In a more advanced stage of the action, during the return phase, additional characterization studies related to aptamer splitting were carried out as preliminary steps towards tuning the binding properties of the aptamer receptors for improving its sensitivity. In parallel, other mechanism previously published to improve sensor sensitivity in a sensing system, known as sequestration, has been successfully achieved and applied in a new aptamer for a specific detection of its target in three different biosensing techniques: EAB sensors (electrochemical read out), ELISA (colorimetric read out) and Lateral Flow assays (optical read out).
In a more advanced stage of the action, during the return phase, additional characterization studies related to aptamer splitting were carried out as preliminary steps towards tuning the binding properties of the aptamer receptors for improving its sensitivity. In parallel, other mechanism previously published to improve sensor sensitivity in a sensing system, known as sequestration, has been successfully achieved and applied in a new aptamer for a specific detection of its target in three different biosensing techniques: EAB sensors (electrochemical read out), ELISA (colorimetric read out) and Lateral Flow assays (optical read out).
Results achieve during the action, both outgoing and incoming phase, comprise the study and exploration of a new labelling architecture for EDNA sensors, the use a novel aptamer to successfully develop a new EAB sensors for chemotherapeutic drug detection, application of a nature inspired mechanism (sequestration) to tune binding properties of receptors in biological assays. Additionally extra efforts were centered in expanding the previous preliminary work to apply other mechanisms (cooperativity) into aptamer sensing systems. With the same aim to incorporate modification to induce more sensitive responses in sensing systems.