Nanoporous interferometers for assessing DogTag/DogCatcher chemistry affinity

This study presents the development of a sensing tool for real-time binding assessment between DogCatcher and DogTag linkers—biochemical functional groups for site-specific protein labeling and conjugation. The system combines nanoporous anodic alumina based Fabry-Pérot interferometers (NAA–FPIs) functionalized with DogCatcher-labelled green fluorescent protein (GFP) and reflectometric interference spectroscopy (RIfS). Protein-modified NAA–FPI platforms were exposed to analytical solutions of GFP-DogTag of different concentrations, from 15 to 100 μg mL –1 . Dynamic changes in the effective optical thickness of protein-modified NAA–FPIs in response to GFP-DogTag were assessed in real time with the objective of evaluating the binding kinetics, affinity, and mechanism. The effective optical thickness of GFP-DogCatcher-modified NAA–FPI structures underwent significant changes upon exposure to GFP-DogTag protein, exhibiting a strong affinity. The selectivity of the system was also assessed by exposing these thin films to analytical solutions of interfering biomolecules. The sensor achieved a sensitivity of 0.56 nm μg mL –1 and a low limit of detection of 9.2 μg mL –1 , with excellent linearity. This system is a promising complement to other detection systems used to assess affinity between biomolecules, expanding our understanding of how this novel class of biomolecules interact in a confined space—with potential implications across a range of biomolecular applications.