Phage Display Affibodies Combined with AuNPs@Ru(bpy)₃²⁺ for Ultra-Sensitive Electrochemiluminescence Detection of Abrin

Abstract

Abrin is a cytotoxin with strong lethality, which is a serious threat to human health and public safety, and thus, highly sensitive detection methods are urgently needed. The phage display affibody has two major modules, among which, the affibody fragment, with small molecular weight, high affinity and easy preparation, can be used for the specific recognition of the target, and the phage shell, with numerous protein copies, can be used as a carrier for the massive enrichment of signal molecules, and thus is particularly suitable as a sensitive probe for signal amplification in high-sensitivity biosensors. In this study, with antibody-coated magnetic microspheres as capture probes, Ru(bpy)32+ and biotin dual-labeled phage display affibodies as the specific signal probes and AuNPs@Ru(bpy)32+ (Ru(bpy)32+-coated gold nanoparticles) as the signal amplification nanomaterials, a new electrochemiluminescence (ECL) biosensor with a four-level sandwich structure of “magnetic capture probe-abrin-phage display affibody-AuNPs@Ru(bpy)32+” was constructed for abrin detection. In this detection mode, AuNPs@Ru(bpy)32+, a gold nanocomposite prepared rapidly via electrical interaction, contained an extremely high density of signal molecules, and the phage display affibodies with powerful loading capacity were not only labeled with Ru(bpy)32+, but also enriched with AuNPs@Ru(bpy)32+ in large amounts. These designs greatly improved the detection capability of the sensor, ultimately achieving the ultra-sensitive detection of abrin. The limit of detection (LOD) was 4.1 fg/mL (3δ/S), and the quantification range was from 5 fg/mL to 5 pg/mL. The sensor had good reproducibility and specificity and performed well in the test of simulated samples. This study expanded the application of affibodies in the field of biosensing and also deeply explored the signal amplification potential of phage display technology, which is of high value for the construction of simple and efficient sensors with high sensitivity.

Keywords

• electrochemiluminescence
• phage display affibody
• gold nanoparticles
• abrin