Antimicrobial drugs are used to control bacterial infections among fish in aquaculture and misuse could be associated with the spread of antimicrobial resistance and tissue residues with a negative impact on humans and animals including fish. This study was conducted to evaluate the effect of cefquinome on Aeromonas hydrophila and some biochemical parameters in tilapia fish, together with estimating its residue in serum, muscles, and liver using high-performance chromatography. One hundred and thirty-five Nile tilapia fish (Oreochromis niloticus) were used and divided into 3 equal groups. Fish of group 1 was a negative control, group 2 was a positive (infected with  Aeromonas hydrophila) and group 3 was infected with the same bacteria and treated with Cefquinome sulphate (Cobactan® 2.5%) as a single intramuscular dose of 10 mg/kg body weight. Cefquinome showed an in-vitro inhibition zone of 29.3± 0.26 mm against A. hydrophilla, and the minimum inhibitory concentration was 2 µg/ ml. While, in-vivo; the mortality rate among infected fish was decreased after treatment by cefquinome from 66.6 % to 17.7 %, a slight recovery has been recorded in biochemical parameters at 7^th to 14^th day post-medication. Serum and tissue residue of cefquinome reached under the limit of detection at 120 hrs. post-medication while still detected in the liver even after 120 hrs post medication. It could be concluded that cefquinome had a good effect against Aeromonas hydrophila infection in tilapia. Fish flesh could be suitable for human consumption 5 days of post-cefquinome  medication.