A quantitative, label-free visual interference colour assay platform for protein targeting and binding assays

Abstract

The vast array of immunoassay technologies used to assess protein interactions is costly or platform-specific. We present a label-free visual interference colour assay (VICA) that quantifies peptide and protein interactions by creating an iridescent surface allowing direct visualisation without spectrophotometric optics or microfluidics. A nanoporous aluminium oxide surface is tuned to match the refractive indices of the overlying protein layers to generate visual interference colours. To functionalise the surface, we created an affinity-capture system using a protein A-carboxyglutamic (GLA) construct that orients antibodies to enhance the signal. Using off-the-shelf antibodies, the platform can isolate analytes in buffer, whole blood, or serum. This surface generates a discernible colour change at concentrations as low as 50 femtomoles/mm2 and can monitor oligomer formation in sequential steps on the same slide. VICA provides comparable kinetic parameters to biolayer interferometry and traditional immunoassays while also allowing characterisation of proteins in large macromolecular complexes.