Figure 1 Preparation procedures of the proposed AFB1 immunosensor for AFB1 detection

 

                  Mycotoxins are widely found in daily products such as peanuts, beans, corn, wheat and rice.  Aflatoxin B1 (AFB1) is one of the most toxic which may affect a serious harm to human and animals, thus causing both acute and chronic diseases.

                  Therefore, a novel sensitive and selective platform for the specific determination of aflatoxin B1 (AFB1) was developed by Associate Prof. Dr. Anchalee Samphao (Department of Chemistry, Faculty of Science, Ubon Ratchathani University).

                   Anti-AFB1 antibody was immobilized on single-walled carbon nanohorns decorated by cobalt oxide composite and gold nanoparticles as shown in Figure 1.  Thereafter, selectivity of the proposed sensor was attributed to the specific binding property of anti-aflatoxin B1 antibody.

                 X-ray crystal structure (PDB entry 5B3N) was used as a structural template for a single-chain fragment variable (scFv) of monoclonal antibody against aflatoxins.  The specific interaction of the proposed immunosensor to AFB1 was clarified using homology modelling combined with molecular docking.  In the presence of AFB1, the current signal of the modified electrode became lower which involved specific antibody-antigen binding (Figure 1), including hydrophobic, hydrogen bonding and pi-pi stack interactions.

                 Close inspection of the predicted structure based on 5B3N revealed that heavy chain (VH) and light chain (VL) contributed to the aflatoxin binding site as shown in Figure 2.  It is found that docked molecule of AFB1 interacts at the surface between the variable domains of both VH and VL (Figure 2), thus contributing to the antigen-binding site.

 

 

Figure 2 The interactions between antibody and aflatoxin B1. The model structure based on 5B3N is shown in cartoon representation whereas heavy chain (VH) is shown in cyan and light chain (VL) is shown in salmon. The labelled amino acid residues are shown as stick models with carbon atoms in cyan (VH) and salmon (VL), nitrogen atoms in blue and oxygen atoms in red. Aflatoxin B1 is shown as a ball-and-stick model with carbon atoms in yellow and oxygen atoms in red. Black dash lines represent hydrogen bonds formed with their distances given in Å. Electrostatic interactions are represented by green dash lines.

                Among four kinds of aflatoxin, it showed that AFB1 has the highest affinity towards antibody, thus considering the binding energy of AFB1.

              The application of the proposed immunosensor in real samples, including peanut, certified reference material of peanut sample (labelled 206 µg kg^-1 AFB1), corn, and chicken feed was investigated.  This present work shows an excellent performance for AFB1 detection which could be applied for food quality control as well as modified to detect other mycotoxins.

 

Re-written by Chomphunuch Songsiriritthigul (Beamline Scientist, SLRI)

 

References

Benkerroum, N. Chronic and acute toxicities of aflatoxins: Mechanisms of action, Int. J. Environ. Res. 17 (2020) 423.

Damphathik, C., Songsiriritthigul, C., Lerdsri, J., Jakmunee, J., Wongnongwa, Y., Jungsuttiwong, S., Ortner, A., Kalcher, K., Samphao, A. A novel immunosensor based on cobalt oxide nanocomposite modified single walled carbon nanohorns for the selective detection of aflatoxin B1. Talanta. (2023). DOI: 10.1016/j.talanta.2023.124472.

Moretti, A., Logrieco, A.F. & Susca, A. Mycotoxins: An Underhand Food Problem, in: A. Moretti, A. Susca (Eds.) Mycotoxigenic Fungi: Methods and Protocols, Springer, New York, New York, NY, 2017, pp. 3-12.