Lateral flow immunoassay for rapid qualitative and quantitative control of the veterinary drug bacitracin in milk

Abstract

The lateral flow method was first developed for the rapid control of the presence and determination of the amount of polypeptide antibiotic bacitracin. The method is based on the competitive interaction of (a) the bacitracin potentially contained in the sample and (b) the bacitracin-protein conjugate immobilized on the membrane for binding with (c) gold nanoparticles labeled by the anti-bacitracin antibodies during the lateral flow of the reagents and the sample along the test strip. The qualitative assessment of the assay results consists of visual control of the presence or absence of bound colored gold nanoparticles in the zone with the immobilized bacitracin-protein conjugate. The quantitative assay uses a photometric measurement of the intensity of this zone coloration. The thresholds for visual and instrumental detection of bacitracin using the developed assay are 100 and 1.0 ng/mL, and the working range of quantitatively measured concentrations is 3 to 80 ng/mL. The average error of measurement is in the range of 1.5%–7.0%. The analysis time is 10 min. The specificity of the assay has been confirmed in relation to other antibiotics. The test system obtains reliable information on the amount of bacitracin in milk without sample preparation. Testing the milk samples (n = 90) showed the coincidence of the results for the developed lateral flow assay and for a commercial enzyme immunoassay kit. The recovery of bacitracin in milk by the proposed lateral flow method ranged from 75 to 140%.