RESEARCH ARTICLE


Development of MNPs Based Enzyme Immuno-Sorbent Analysis for the Determination of Organophosphorus Pesticides in Milk



Yavor Ivanov1, *
1 Department of Biotechnology, “Prof. Dr Assen Zlatarov” University, Burgas, Bulgaria


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Creative Commons License
© 2019 Yavor Ivanov.

open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0), a copy of which is available at: (https://creativecommons.org/licenses/by/4.0/legalcode). This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

* Address correspondence to this author at the Department of Biotechnology, “Prof. Dr Assen Zlatarov” University, Burgas, Bulgaria;
E-mail: qvor_burgas@abv.bg


Abstract

Objectives:

High toxicity of pesticides requires accurate and reliable methods to monitor their levels for food, water and soil safety.

Methods:

Magnetic nanoparticles were synthesized and functionalized with (3-aminopropyl)triethoxysilane. Sheep polyclonal-anti-paraoxon and anti-dichlorvos antibodies were produced. Dichlorvos-cationized ovalbumin-HPR and paraoxon-ovalbumin-HPR conjugates were prepared and characterized. The optimal conditions for antibody immobilization were determined. Enzyme immunosorbent assay based on Magnetic Nanoparticles (MNPs) for the determination of pesticides was developed. A comparison of the developed modified MNPs-based ELISA with the conventional ELISA was performed. The effect of protein, pH, and fats in milk samples on the analytical characteristics of the MNPs-based ELISA was investigated. In order to validate the results obtained from the MNPs-based ELISA method, experiments with HPLC were performed.

Results:

The linear range of the standard curve for the determination of paraoxon in buffer with anti-paraoxon antibody was 0.125-5 ng/mL, for the dichlorvos with anti-dichlorvos antibody was 0.0625-5 ng/mL, and for the mixture of paraoxon and dichlorvos in ratio (1:1) with multi-antibody was 0.125-5 ng/mL. A comparison of the developed modified MNPs-based ELISA with the conventional ELISA was performed. The recovery of the obtained results for the determination of pesticides in milk samples in both methods was from 104 to 110%.

Conclusion:

The stated advantages of magnetic nanoparticles provide good sensitivity and rapidity of immunoassay and a simple procedure for separating of antibody-magnetic nanoparticles with the bound antigen from the non-bound antigen in the sample.

Keywords: Pesticides, Polyclonal antibody, Magnetic nanoparticles, Enzyme immuno-sorbent analysis, Milk, ELISA method.