Specific and reversible immobilization of proteins tagged to the affinity polypeptide C-LytA on functionalized graphite electrodes

We have developed a general method for the specific and reversible immobilization of proteins fused to the choline-binding module C-LytA on functionalized graphite electrodes. Graphite electrode surfaces were modified by diazonium chemistry to introduce carboxylic groups that were subsequently used to anchor mixed self-assembled monolayers consisting of N,N-diethylethylenediamine groups, acting as choline analogs, and ethanolamine groups as spacers. The ability of the prepared electrodes to specifically bind C-LytA-tagged recombinant proteins was tested with a C-LytA-β-galactosidase fusion protein. The binding, activity and stability of the immobilized protein was evaluated by electrochemically monitoring the formation of an electroactive product in the enzymatic hydrolysis of the synthetic substrate 4-aminophenyl β-D-galactopyranoside. The hybrid protein was immobilized in an specific and reversible way, while retaining the catalytic activity. Moreover, these functionalized electrodes were shown to be highly stable and reusable. The method developed here can be envisaged as a general, immobilization procedure on the protein biosensor field.

Methodology based on registration techniques for representing subjects and their deformations acquired from general purpose 3D sensors

In this thesis a methodology for representing 3D subjects and their deformations in adverse situations is studied. The study is focused in providing methods based on registration techniques to improve the data in situations where the sensor is working in the limit of its sensitivity. In order to do this, it is proposed two methods to overcome the problems which can difficult the process in these conditions. First a rigid registration based on model registration is presented, where the model of 3D planar markers is used. This model is estimated using a proposed method which improves its quality by taking into account prior knowledge of the marker. To study the deformations, it is proposed a framework to combine multiple spaces in a non-rigid registration technique. This proposal improves the quality of the alignment with a more robust matching process that makes use of all available input data. Moreover, this framework allows the registration of multiple spaces simultaneously providing a more general technique. Concretely, it is instantiated using colour and location in the matching process for 3D location registration.