Gold nanoparticle-based localized surface plasmon immunosensor for staphylococcal enterotoxin A (SEA) detection
We describe the engineering of stable gold nanoparticle (AuNP) bioconjugates for the detection of staphylococcal enterotoxin A (SEA) using localized surface plasmon resonance (LSPR). Two types of AuNP bioconjugates were prepared by covalently attaching anti-SEA antibody (Ab) or SEA to AuNPs. This was achieved by reacting Traut’s reagent with lysine residues of both proteins to generate thiol groups that bind to gold atoms on the AuNP surface. These bioconjugates were characterized in-depth by absorption spectroscopy, cryo-transmission electron microscopy, dynamic light scattering, and zeta potential measurements. Their stability over time was assessed after 1 year storage in the refrigerator at 4 °C. Two formats of homogeneous binding assays were set up on the basis of monitoring of LSPR peak shifts resulting from the immunological reaction between the (i) immobilized antibody and free SEA, the direct assay, or (ii) immobilized SEA and free antibody, the competitive assay. In both formats, a correlation between the LSPR band shift and SEA concentration could be established. Though the competitive format did not meet the expected analytical performance, the direct format, the implementation of which was very simple, afforded a specific and sensitive response within a broad dynamic range—nanogram per milliliter to microgram per milliliter. The limit of detection (LOD) of SEA was estimated to equal 5 ng/mL, which was substantially lower than the LOD obtained using a quartz crystal microbalance. Moreover, the analytical performance of AuNP-Ab bioconjugate was preserved after 1 year of storage at 4 °C. Finally, the LSPR biosensor was successfully applied to the detection of SEA in milk samples. The homogeneous nanoplasmonic immunosensor described herein provides an attractive alternative for stable and reliable detection of SEA in the nanogram per milliliter range and offers a promising avenue for rapid, easy to implement, and sensitive biotoxin detection.
KeywordsImmunosensor Localized surface plasmon resonance Staphylococcal enterotoxin A Gold nanoparticles
We would like to thank the DIM Analytics and Region Ile-de-France for M. Ben Haddada PhD scholarship. We also thank Anton Paar for the access to Litesizer™ 500 apparatus. This work was supported by the iFood initiative Nanyang Technological University, by the French-Singaporean PHC Merlion program (grant 5.03.15), and by the ANR-FWF program (grant ANR-15-CE29-0026).
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Conflict of interest
The authors declare that they have no competing interests.
- 2.Le Loir Y, Baron F, Gautier M. Staphylococcus aureus and food poisoning. Genet Mol Res. 2003;2(1):63–76.Google Scholar
- 9.Mayer KM, Lee S, Liao H, Rostro BC, Fuentes A, Scully PT, et al. A label-free immunoassay based upon localized surface plasmon resonance of gold Nanorods. ACS Nano. 2008;2(4):687–92. doi: 10.1021/nn7003734.
- 11.Liu XH, Wang Y, Chen P, Wang YS, Mang JL, Aili D, et al. Biofunctionalized gold nanoparticles for colorimetric sensing of Botulinum neurotoxin A light chain. Anal Chem. 2014;86(5):2345–52. doi: 10.1021/ac402626g.
- 13.Montenegro J-M, Grazu V, Sukhanova A, Agarwal S, de la Fuente JM, Nabiev I, et al. Controlled antibody/(bio-) conjugation of inorganic nanoparticles for targeted delivery. Adv Drug Deliv Rev. 2013;65(5):677–88. doi: 10.1016/j.addr.2012.12.003.
- 19.Ben Haddada M, Huebner M, Casale S, Knopp D, Niessner R, Salmain M, et al. Gold nanoparticles assembly on silicon and gold surfaces: mechanism, stability, and efficiency in diclofenac biosensing. J Phys Chem C. 2016;120:29302–11.Google Scholar
- 23.Hermanson GT. Chapter 24—Preparation of colloidal gold-labeled proteins. In: Hermanson GT, editor. Bioconjugate techniques. Second ed. New York: Academic Press; 2007. p. 924–35. doi: 10.1016/B978-0-12-370501-3.00024-2.
- 27.Thobhani S, Attree S, Boyd R, Kumarswami N, Noble J, Szymanski M, et al. Bioconjugation and characterisation of gold colloid-labelled proteins. J Immunol Methods. 2010;356(1–2):60–9. doi: 10.1016/j.jim.2010.02.007.
- 35.Salmain M, Ghasemi M, Boujday S, Spadavecchia J, Techer C, Val F, et al. Piezoelectric immunosensor for direct and rapid detection of staphylococcal enterotoxin A (SEA) at the ng level. Biosens Bioelectron. 2011;29(1):140–4. doi: 10.1016/j.bios.2011.08.007.