Immunofluorescent Analysis with Magnetic Nanoparticles for Simultaneous Determination of Antibiotic Residues in Milk

Abstract

An analytical system for simultaneous detection of three antibiotic residues in milk (penicillin, tetracycline, and sulfadimethoxine sodium salt) was developed. The method was based on a fluorescence immunoassay. Magnetic nanoparticles (MNPs) were prepared and then coated with 3-(aminopropyl) triethoxysilane (APTES). Antibodies, against penicillin, tetracycline, and sulfadimethoxine, were immobilized on APTES-MNPs. Two immobilization methods were used: a random and an oriented method by protein A. The fluorochrome-antigen conjugates, containing FITC, ATTO 590, and ATTO 630 were obtained. Three separate immunofluorescent analyses were investigated for penicillin, tetracycline, and sulfadimethoxine, first in buffer and then in milk. Linearity values of standard curves in milk were: for penicillin 4–15 ng mL⁻¹, for tetracycline 50–500 ng mL⁻¹ (using random immobilization) and 100–500 ng mL⁻¹(using oriented immobilization), and for sulfadimethoxine 100–500 ng mL⁻¹. The linearity and sensitivity of calibration curves of determination of penicillin, tetracycline and sulfadimethoxine in milk were very close to the obtained results for separate determination of three antibiotics. It was shown that the multiple immunofluorescence analysis for antibiotic residues in milk without preliminary treatment is effective and rapid.

Keywords:

• Antibiotics
• Fluorescence immunoassay
• Magnetic nanoparticles
• Milk
• Multiple analysis