Quantum Dot–Based FRET Immunoassay for HER2 Using Ultrasmall Affinity Proteins
Abstract
Engineered scaffold affinity proteins are used in many biological applications with the aim of replacing natural antibodies. Although their very small sizes are beneficial for multivalent nanoparticle conjugation and efficient Förster resonance energy transfer (FRET), the application of engineered affinity proteins in such nanobiosensing formats has been largely neglected. Here, it is shown that very small (≈6.5 kDa) histidine‐tagged albumin‐binding domain‐derived affinity proteins (ADAPTs) can efficiently self‐assemble to zwitterionic ligand–coated quantum dots (QDs). These ADAPT–QD conjugates are significantly smaller than QD‐conjugates based on IgG, Fab', or single‐domain antibodies. Immediate applicability by the quantification of the human epidermal growth factor receptor 2 (HER2) in serum‐containing samples using time‐gated Tb‐to‐QD FRET detection on the clinical benchtop immunoassay analyzer KRYPTOR is demonstrated here. Limits of detection down to 40 × 10−12m (≈8 ng mL−1) are in a relevant clinical concentration range and outperform previously tested assays with antibodies, antibody fragments, and nanobodies.
Document Details
- Document Type
- Pub Defense Publication
- Publication Date
- Aug 01, 2018
- Source ID
- 10.1002/smll.201802266
Entities
People
- Igor L. Medintz
- Kimihiro Susumu
- Niko Hildebrandt
- Sarah Lindbo
- Sophia Hober
- Xue Qiu
- Yu‐tang Wu
Organizations
- Agence Nationale de la Recherche
- Horizon 2020
- Institut Universitaire de France
- KeyW Corporation
- Office of Naval Research
- Royal Institute of Technology
- United States Naval Research Laboratory