The Open Biotechnology Journal

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Simultaneous Determination of Penicillin G and Chloramphenicol in Milk by a Magnetic Nanoparticle-Based Fluorescent Immunoassay

Milka Atanasova1, Yavor Ivanov1, Elena Zvereva2, Anatoly Zherdev2, Tzonka Godjevargova1, *
1 Department of Biotechnology, “Prof. Dr Asen Zlatarov” University, Burgas, Bulgaria
2 A.N. Bach Institute of Biochemistry, Research Centre of Biotechnology of the Russian Academy of Sciences, Moscow, Russia



Antibiotic residues are a problem of increasing importance and have direct consequences for human and animal health. The frequent use of antibiotics in veterinary practice causes their excretion in milk in dairy cattle. This way, they can easily enter the human body through the consumption of milk and dairy products.


This induces the need for accurate and sensitive methods to monitor antibiotic levels in milk. The aim of this study was to develop a rapid and sensitive magnetic nanoparticle-based fluorescence immunoassay for the simultaneous detection of chloramphenicol and penicillin G in milk.


Magnetic nanoparticles were synthesized and functionalized with (3-aminopropyl)triethoxysilane. Chloramphenicol-Ovalbumin and Chloramphenicol-Ovalbumin-Fluorescein-5-isothiocyanate conjugates were prepared. Penicillin G – ATTO 633 fluorescent conjugate was synthesized. Antibodies against chloramphenicol and penicillin G were immobilized onto the magnetic nanoparticles. The competitive fluorescent immunoassay was developed. The optimal concentration of the antibody-magnetic nanoparticles and the fluorescent conjugates for the assay was determined. The calibration curves for the antibiotics in buffer and milk were plotted. Fluorescent immunoassay for the simultaneous determination of chloramphenicol and penicillin G in milk was developed.


The limit of detection by the simultaneous immunoassay of chloramphenicol and penicillin G in milk was 0.85 ng/mL and 1.6 ng/mL, respectively. The recovery of different concentrations of chloramphenicol and penicillin G in milk samples varied from 98% to 106%.


A rapid and sensitive magnetic nanoparticle-based immunofluorescent assay for the simultaneous determination of chloramphenicol and penicillin G in milk was developed. The magnetic nanoparticles ensured rapid and easy procedure.

Keywords: Chloramphenicol, Penicillin G, Milk, Magnetic nanoparticles, Fluorescent immunoassay, Antibiotic.

Article Information

Identifiers and Pagination:

Year: 2020
Volume: 14
First Page: 59
Last Page: 69
Publisher Id: TOBIOTJ-14-59
DOI: 10.2174/1874070702014010059

Article History:

Received Date: 31/12/2019
Revision Received Date: 14/02/2020
Acceptance Date: 03/03/2020
Electronic publication date: 16/06/2020
Collection year: 2020

© 2020 Atanasova et al.

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 Asen Zlatarov” University, Burgas, Bulgaria; E-mail: godjevargova@

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