Single-Step Homogeneous Immunoassays Utilizing Epitope-Tagged Gold Nanoparticles: On the Mechanism, Feasibility, and Limitations

Abstract

A single-step gold nanoparticle (AuNP)-based immunoassay is demonstrated in which the nanoparticle surface is tagged with short viral peptide epitopes. Antiviral antibodies with monoclonal specificity trigger nanoparticle aggregation yielding a colorimetric response that enables detection of antibodies in the low-nanomolar range within a few minutes. In silico insights into the interactions at the epitope−gold interface demonstrate that the conformational landscape exhibited by the epitopes is strongly influenced by the amino acid sequence and location of particular residues within the peptides. The conformation, orientation, and linker chemistry of the peptides affect the immune complex formation in nonintuitive ways that are, nevertheless, explained by a unique sterically kinetically driven aggregation mechanism. The rapid and specific performance of the AuNP immunoassay may have generic potential in point of care diagnostics and other laboratory routines.

SERS immunoassay based on the capture and concentration of antigen-assembled gold nanoparticles

“…2, the mean hydrodynamic diameter increased after introduction of the target mouse IgG antigen, reaching a maximum size increase of ∼100 nm after 90 min of incubation. These aggregation kinetics are consistent with other groups reporting that 80% of aggregation is completed in ∼60 min [25]. Importantly, the antibody-antigen binding time in this homogeneous format significantly outperforms that reported for heterogeneous binding and is approaching practical timescales for point-of-care diagnostic assays [26,27].…”

Gold Nanoparticles for In Vitro Diagnostics

“…The basis of color change is that the interparticle spacing of the cross-linked AuNPs should be smaller than the diameter of the nanoparticles. 84 To break this barrier, Stevens and co-workers 85 used 60 nm AuNPs, whose surfaces were functionalized with short viral peptide epitope, to detect antiviral epitope antibodies. By adding the analytes, significant changes in the absorption band as well as colorimetric response were yielded, most likely due to the fact that the interparticle spacing of the particle networks was greatly reduced.…”

Understanding and Designing the Gold–Bio Interface: Insights from Simulations

“…The interplay between sequence, conformation(s) and binding propensity has a significant impact on the affinity of peptides to gold. For example, some peptide residues can show variability in their degree of surface interaction depending on their position in the chain and their immediate environment and this requires careful consideration …”

“…Copyright 2013, American Chemical Society and c) epitope‐tagged AuNP biosensors. Adapted and reproduced with permission . Copyright 2014, American Chemical Society.…”