Thiol-ene coupling (TEC) and Light Assisted Molecular Immobilization (LAMI) are photonic techniques leading to immobilization of bioreceptors, such as, antibodies which recognize cardiac markers. These techniques present advantages compared to traditional immobilization techniques since, e.g., there are no thermal or chemical steps and they work in water media. TEC reaction takes place at close-to-visible wavelengths (λ=365nm) which induces the formation of thiol radicals which bind to alkene functional group on the surface through a thioether bond. LAMI secures molecular immobilizations in a spatially oriented, localized and covalent coupling of biomolecules onto thiol reactive surfaces down to submicrometer spatial resolution. LAMI is possible due to a conserved structural motif in proteins: the spatial proximity between aromatic residues and disulfide bridges. When aromatic residues are excited with UV light (275- 295nm), disulphide bridges are disrupted and free thiol groups are formed that can bind covalently to a surface decorated with thiol groups.
We have achieved successful immobilization of anti-troponin and anti-myoglobin antibodies with both photonic immobilization techniques. The microarrays of immobilized monoclonal antibodies have successfully detected the CVD biomarkers troponin I and myoglobin, as confirmed by fluorescence imaging. A sandwich immunoassay was carried out, Troponin I and Myoglobin were detected down to 10 ng/mL and 1 ng/mL, respectively.