Antibiotics such as the quinolones, tetracycline, streptomycin and lincomycin are increasingly found in dairy products. To ensure consumer safety, a rapid screening method is needed that is accurate and cost-effective. We tested a multiplexed detection technology for quantification of these 4 antibiotics simultaneously in milk. The method relies on an enzyme linked immunosorbent assay-based visualization microarray chip technique that can detect multiple target molecules simultaneously. The microarray chip was fabricated by adsorption of BSA- or OVA-conjugated small molecules as haptens on the microplates. Using competitive immunoassay, combined with silver nanoparticle (AgNP) catalyzed visualization technology to quantify multiple substances in milk at the same time. This colorimetric technology provides direct results that can be detected using common commercial microplate scanners. In this study, the LOD of the four different antibiotics were calculated by determining the mean response of 20 negative milk samples and adding 3 times the standard deviation. The LOD was defined at 3.30, 3.39, 2.42, and 4.88 μg kg⁻¹ for quinolones, tetracycline, lincomycin and streptomycin, respectively, the LOQ values were 8.97, 8.43, 5.36 and 10.97 μg kg⁻¹ for quinolones, tetracycline, lincomycin and streptomycin, respectively. These limits are less than the maximum residue levels established in China, the Codex Alimentarius Commission, and the European Union. The accuracy of method was also adequate for these four substances and experiments using actual samples demonstrated recovery rates ranging from 77.6 to 116.4%. The 198 liquid milk samples were analyzed using this method. This technology could achieve the simultaneous detection of analytes and will have broad applications in the food safety analysis field.