Aim: The present study was described thin layer chromatography (TLC) and ultra-high performance liquid chromatography (UHPLC) method for the detection of antibacterial substances in poultry muscle (breast and thigh), kidney, and liver. Materials and Methods: TLC method was used for screening detection of tetracycline, amoxicillin, ciprofloxacin, and enrofloxacin residues in poultry tissues. The samples were extracted with trichloroacetic acid (30%), diethyl ether, followed by detection in pre-coated TLC paper on ultraviolet detector. The UHPLC method was used for the quantification of antimicrobial residues in poultry tissues. Results: The residues of tetracycline were 48% in livers, 24% in kidneys, 20% in thigh muscles, and 24% in breast muscles. Ciprofloxacin residues were found 44% in liver, 42% in kidneys, 34% in thigh muscles and 30% in breast muscles. Enrofloxacin residues were found 40% in liver, 34% in kidneys, 22% in thigh muscles, and 18% in breast muscles. Amoxicillin residues were found 42% in liver, 30% in kidneys, 26% in thigh muscles and 22% in breast muscles. Most of the cases highest residues were found in liver such as tetracycline (48%), ciprofloxacin (44%), enrofloxacin (40%) and amoxicillin (42%) and almost lowest in breast muscles. In addition, nine positive samples from broiler were selected for amoxicillin residue quantification by UHPLC. It was observed that the concentration of amoxicillin residue in liver was ranging from 16.92 μg/kg to 152.62 μg/kg and in breast muscle was 45.38 μg/kg to 60.55 μg/kg, respectively. The maximum and minimum peak time was 4.7-5.2 min. Among the poultry tissues, liver had the highest level of antibiotic residues in comparison to other samples but the variation was not significant (p>0.05). Conclusions: Evidence suggests that more judicious use of antimicrobials in food animals will reduce the selection of resistant bacteria and help to preserve these valuable drugs for both human and veterinary medicine. The method described in this study is a simple, easy inexpensive which can be readily adopted by any laboratory for the detection of antibiotic residues in tissues of food-producing animals.