Nano-Magnetic Immunosensor Based on Staphylococcus Protein A and the Amplification Effect of HRP-Conjugated Phage Antibody

Abstract

In this research, super-paramagnetic Fe3O4 nanoparticles (magnetic particles) were coated with Staphylococcus protein A (SPA) and coupled with polyclonal antibody (pcAb) to construct magnetic capturing probes, and HRP-conjugated phage antibody was then used as specific detecting probe to design a labeled immunosensor for trace detection of Staphylococcus aureus enterotoxin B (SEB). The linear detection range of the sensor was 0.008~125 µg/L, the regression equation was Y = 0.487X + 1.2 (R = 0.996, N = 15, p < 0.0001), the limit of detection (LOD) was 0.008 µg/L, and the limit of quantification (LOQ) was 0.008 µg/L. HRP-conjugated phage antibody, SPA and magnetic particles can enhance the sensitivity 4-fold, 3-fold and 2.6-fold higher, respectively. Compared with conventional double-antibody sandwich ELISA, the detection sensitivity of the sensor was 31-fold higher resulting from the integrated amplifying effect. The immunosensor integrates the unique advantages of SPA-oriented antibody as magnetic capturing probe, HRP-conjugated phage antibody as detecting probe, magnetic separation immunoassay technique, and several other advanced techniques, so it achieves high sensitivity, specificity and interference-resistance. It is proven to be well suited for analysis of trace SEB in various environmental samples with high recovery rate and reproducibility.

Keywords

• Staphylococcus protein A
• HRP-conjugated phage antibody
• signal amplification
• nanomagnetic immunosensor
• Staphylococcus aureus enterotoxin B