The uptake of proteomics for precision or personalised medicine applications requires the development of an effective screening instrument capable of detecting proteins at the low concentrations they are found in biological conditions while addressing their large chemical variability.
The ElectroMed project's objective is to build and validate a proof-of-concept prototype of a programmable high-throughput peptide microarray technology. It is based on the radical vision of integrating electrochemical synthesis of peptide bioreceptors to enable programmable in situ protein detection with nano-functionalised FinFET sensors for high-performance data acquisition within a microfluidic-driven multiplexed platform for parallel screening.
The project will undertake the research and innovation work required to
integrate EPS and the FinFET sensors in a single cost-effective prototype that we will validate in laboratory-relevant conditions.
The ElectroMed project will achieve a technology breakthrough of the first fully-programmable in situ protein screening instrument. This prototype small-size single instrument will come as a faster, cheaper, and more efficient technology than current protein screening instruments.
The potential impact is to significantly ease and reduce the price of protein screening to trigger an uptake of protein detection in healthcare (our primary focus) as well as in the food sector (food authenticity and traceability, allergens, GMOs or toxins detection all along the food value chain), environmental remediation or defence (detection of biological warfare agents).
A User guide on sequential injection to make the sequential injection system more user-friendly and improve the usability was developed within the frame of ElectroMed and is accessible on demand on Elveflow's website.
Sensors developed in Twente University in collaboration with LIST consist of FET devices compatible with wafer scale fabrication.
Acid labile peptide synthesis for semiconductor oxides .
Semiconductor oxides and metal oxides represent an important class of substrates to detect peptide-protein interactions using biosensors. Silicon dioxide (glass) is probably the most accessible substrate as it is the main component of microscope slides. Techniques such micro spotting use these substrates to immobilize peptide sequences to detect then the interactions with proteins by fluorescence. Silicon-dioxide and metal oxides such as Hf2O3 or TiO2 areused as the interface of biosensors such as field effect transistors (FET), which can be used to detect the interactions of peptides with proteins with lower quantities than fluorescence using miniaturized biosensors. In ElectroMed we have developed a method able to locally produce acidity, which can be integrated with FET sensors that use a semiconductor oxide or a metal oxide interface between the liquid medium and the solid-state transducer. This is why we developed a compatible acid labile solid-phase synthesis.
We three different classes of amino acids attending to its complexity for the synthesis:
Bifunctional amino acids (AAs), in our study represented by valine, leucine and isoleucine.
Trifunctional protected amino acids, represented by lysine, aspartic acid and glutamic acid.
Trifunctional non-protected amino acids, where we used serine, tyrosine, tryptophan and histidine and threonine.
We resolved a base for a proof of concept with a limited number of AAs:
-We studied bifunctional and trifunctional (E and D) AAs obtaining ≥90% purity (crude peptides without purification).
-We studied different solvents, including green solvents. We concluded that Acetonitrile can be used as a solvent, which is the safest possibility in our system.
-We tested the deprotection of acid labile groups with Trifluoroacetic Acid as well as Hydrochloric Acid, both obtaining high yields on the control of the synthesis.
-We obtained high purities for sequences with protected Amino Acids, and the possibilities to synthetise some non -protected Amino Acids with non protected sequences.
-We studied the synthesis of longer sequences (octa/decapeptide) ≥90% purity
-We optimised the synthesis of unprotected hydroxy AAs (S, T, Y), which is still a challenge. However, we were able to produce samples with ≥81% purity
-We completed and optimized the list of bifunctional Amino Acids (adding L, I and V): ≥93% purity
-We used Control Porous Glass also with W, Q and N amino Acids:
a) W and Q: ≥96% purity
b) N: ≥80% purity
-Currently, R and H remain as AAs with no base labile protecting groups are available.
- DYK peptides were synthesized with moderate results: 57-75% purity