UCPH first investigated and assessed the relevance of linear peptide sequences that can interact with the immune system. The Cytomegalovirus (HHV-5) proteome contains peptides that interact with human antibodies and MHC Class II molecules, resembling cancer-related neopitopes. Furthermore, in collaboration with University Hospital Copenhagen, UCPH collected serum samples from COVID-19 and non-COVID-19 patients. Novel peptide epitope targets were identified and selected for single residue mapping; tested for inhibition of neutralizing anti-SARS-CoV2 antibodies; and used to investigate the presence of peptide-specific antibodies in a cohort of Covid-19 patients.
ElectroMed scientists, led by ELV, developed a functional prototype for sequential injection of multiple solutions using an Ibidi chip. The system has overcome the challenges of working with organic chemistry to miniaturize liquid management, resulting in a fully atomized system capable of delivering parallel peptide synthesis at a rate of one amino acid every five minutes, with the potential to reduce the synthesis time to less than a minute for each amino acid addition. Also, liquid volumes have been reduced, and in our current protocol, we use half ml with 0.3M concentrations, with the possibility of further reducing concentrations and dead volumes.
The synthesis takes place in a microfluidic chamber designed by LIST, where electrochemically generated acid is produced in addressable FET sensors that can monitor the synthesis process. We demonstrated the electrochemical production of acid quantities equivalent to traditional synthesis, which can be used to deprotect acid labile groups such as BOC in each addressable sensor. Furthermore, we completed the synthesis of the first peptide, resulting in a three-amino acid sequence detected by an antibody interaction, which was validated throughout our project.
UPF developed a protocol for coupling standard amino acids on surfaces similar to those used in sensors. This involved using APTES functionalized porous glass beads with adequate yield for HPLC-MS. The protocol was validated in collaboration with LIST and ELV in planar surfaces using mass spectrometry and SPR, allowing for continuous monitoring of the synthesis process.
The response of our FET sensors was modelled through a collaboration between LIST and UoT, led by UoG. We developed a surface interface model that can be used to predict the sensor response to peptide protein interactions as well as to provide a chemical characterisation of the sensor interface that can be used to monitor the synthesis and provide non-destructive quality control of each spot, which is a unique feature of our technology. In addition, we discovered a method for detecting amino acid and peptide fingerprints using the sensor signal from our FET.
UoT created FET sensors based on planar junctionless transistors, which can provide greater reliability. Our initial studies demonstrated that we can detect the chemical fingerprints of changes occurring during peptide synthesis using the detection of Boc protection and deprotection. To accomplish this, UoT created a control system that can accurately control pH levels.
