Preparation of IgY Oriented Conjugated Fe3O4 MNPs as Immunomagnetic Nanoprobe for Increasing Enrichment Efficiency of Staphylococcus aureus Based on Adjusting the pH of the Solution System

Abstract

Immunomagnetic separation based on Fe₃O₄ magnetic nanoparticles (MNPs) has been widely performed in sample pretreatment. The oriented conjugation strategy can achieve a better capture effect than the N-(3-dimethylamlnopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) /N-hydroxysuccinimide (NHS) method. However, immunoglobulin yolk (IgY) cannot be oriented through an SPA strategy like immunoglobulin G (IgG). In this article, an oriented conjugation nanoprobe was prepared for the enrichment of bacteria based on pH adjusting. The main factors affecting the enrichment efficiency were studied, such as the pH of the buffer system, the concentration of IgY, the concentration of nanoprobe, and the enrichment time. Under the optimal conditions, the enrichment efficiency toward target bacteria could reach 92.8%. Combined with PCR, the limit of detection (LOD) was found to be 10³ CFU/ml, which was lower than the PCR only. In conclusion, we provided a new protocol for the oriented conjugation of IgY and high sensitivity detection with simple pretreatment.

Keywords: Fe3O4 magnetic nanoparticles (MNPs); Staphylococcus aureus; immunoglobulin yolk (IgY); immunomagnetic separation; oriented conjugation.