Background:In an attempt to enhance the quality and quantity of food production (especially milk) and in order to prevent, or treat,animal diseases, the use of antibiotics in Algeria follows an increasing trend. The increased use evidently contributes to the emergence of increased contamination levels of antibiotic residues. Results: In this work, two methods were used to detect presence of antibiotic residues in raw and fermented cow's milk collected in Guelma's farms (in Algeria). The screening comprised different points of sale in Guelma province. In a first step a widely used prescreening method based on microbial inhibition assay; Delvotest SP-NT; was used to analyze 131 milk samples. In a second step a liquid chromatography coupled to mass spectrometry (LC-MS/MS) was used. The latter was first optimized for extraction of 36 veterinary drugs of penicillins, quinolones, macrolides, tetracyclines, sulfonamides, and trimethoprim from the collected milk. After simple extraction and dilution, the 194 samples, including those previously tested by the Delvotest SP-NT, were analyzed by LC-MS/MS. Results obtained by both methods were compared. Among the LC-MS/MS findings, 65.46 % of non-conform samples contained authorized residues at levels higher than the MRL, residues without set MRL, or non-authorized residues.Conclusion: The comparison of both methods showed that Delvotest SP-NT is less trustworthy due to number of false negative results. This was further confirmed by LC-MS/MS pointing out the traces of antibiotics in numerous samples. In 65.46 % of milk samples residues of antibiotics were found suggesting a lack of public health controls as well as an evidence of the negligent use of antibiotics in the livestock industry, which both form a risk to public health. This indication should be confirmed by a nationwide study with in-depth analyses of antibiotic's presence in food chain originated from animals. The study offered an LC-MS/MS based analytical method ready to be used in Algerian National Residues Control Plan as a versatile analytical tool to monitor and determine the occurrence of antibiotic multi-residues in milk.