A Dual-Color Quantum Dots Encoded Frit-Based Immunoassay for Visual Detection of Aflatoxin M₁ and Pirlimycin Residues in Milk

Mycotoxins and antibacterial agents are the main chemical hazards that lead to several health problems. Nowadays, multiplex immunoassay is a primary goal throughout the world. Here, aflatoxin M₁ and pirlimycin were selected as models, and a novel dual colorimetric encoded frit-based immunoassay was developed for simultaneously screening of aflatoxin M₁ and pirlimycin residues in milk. This multiplex frit-based immunoassay combined two monoclonal antibodies to extend the spectrum of analytes and to enable detection of two classes of analytes in a single test. The cutoff values were 0.02 μg/kg for aflatoxin M₁ and 0.5 μg/kg for pirlimycin, which satisfied the requirement to measure the maximum residue levels. The novel colorimetric frit-based immunoassay has the advantage of high throughput, short analysis time, reduced overall cost per assay, and can be used as a rapid screening technique for simultaneously detecting aflatoxin M₁ and pirlimycin residues in milk

Analysis of Pirlimycin Residues in Beef Muscle, Milk, and Honey by a Biotin–Streptavidin-Amplified Enzyme-Linked Immunosorbent Assay

Food contamination by veterinary drug residues is a worldwide public health concern and requires continuous monitoring. In this study, we developed a biotin–streptavidin-amplified ELISA (BA-ELISA) using a produced monoclonal antibody for detecting pirlimycin residues in beef muscle, milk, and honey. The IC₅₀ value of the BA-ELISA was 1.6 ng/mL for pirlimycin in buffer, and the sensitivity was improved 3 times compared to traditional ELISAs. The optimized BA-ELISA can be used to quantitate trace amounts of pirlimycin residues in beef muscle, milk, and honey. This method had limits of detection (LODs) of 4.45 μg/kg in beef muscle, 1.65 μg/L in milk, and 2.75 μg/kg in honey. The average recovery of the BA-ELISA ranged from 78 to 97%, and the coefficient of variation ranged from 5.3 to 13.5%. The developed BA-ELISA method was validated using LC-MS/MS, and the BA-ELISA can be used for routine screening analysis of pirlimycin residues