A preliminary screening assay based on a microbial chromogenic reaction was developed to detect common antibiotic residues in meat rapidly. The assay comprised two bioassays: one for Escherichia coli and another for Geobacillus stearothermophilus. The assay was optimized and evaluated for the simultaneous screening of 30 antibiotics from five common antibiotic classes (tetracyclines, aminoglycosides, macrolides, beta-lactams, and quinolones) found in meat. Extraction using phosphate-acetonitrile buffer (pH 7.2) and a delipidating treatment using n-hexane resulted in a high extraction efficacy for the five antibiotics, without affecting the microbial color reaction. A carrier, polyvinyl alcohol (0.1 g/mL); a cross-linking agent, boric acid-sodium tetraborate solution (pH 5.5); and a bacterial suspension with an initial optical density of 1.0 were the optimal embedding conditions for stability, microbial activity, and chromogenic efficiency. The assay exhibited a 6-month shelf life, with detection limits of 40-60, 60-140, 60-100, 20-40, and 40-180 mu g/kg for tetracyclines, aminoglycosides, macrolides, beta-lactams, and quinolones, respectively, which met the European Commission (37/2010) requirements for antibiotic residue limits. Our assay results were confirmed using LC-MS/MS with 160 samples, revealing a good correlation. This study demonstrates a reliable, easy-to-use, and economical method for preliminary screening of antibiotic residues in meat. This method may find an immediate application in food safety and general testing laboratories.