The Open Biotechnology Journal

ISSN: 1874-0707 ― Volume 14, 2020

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



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


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.


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%.


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.

Article Information

Identifiers and Pagination:

Year: 2019
Volume: 13
Issue: Suppl-1, M6
First Page: 146
Last Page: 154
Publisher Id: TOBIOTJ-13-146
DOI: 10.2174/187407070190130146

Article History:

Received Date: 25/04/2019
Revision Received Date: 24/07/2019
Acceptance Date: 13/10/2019
Electronic publication date: 15/11/2019
Collection year: 2019

© 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: ( 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;

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