In this study, a water-soluble fluorescent probe based on copper nanoparticles (CuNPs) was synthesized using a hydrothermal method with bovine serum albumin (BSA) (BSA@CuNPs) as a protectant, and it was utilized to efficiently detect the concentration of tetracycline (TC). Upon the addition of TC to the BSA@CuNPs probe, its fluorescence intensity is obviously diminished due to the keto-enol functions C=O group in TC connects with the hydroxyl group on the BSA@CuNPs probe to form a non-fluorescent complex, which can be successfully utilized for detecting TC concentration. Linear range and detection limit for TC detection are 0.1—130 μM and 90 nM. Furthermore, the BSA@CuNPs probe was applied to detect TC concentrations in real meat samples, with recoveries ranging from 95.40% to 102.13%, and the relative standard deviation (RSD) was below 2.73%. Therefore, BSA@CuNPs probe exhibits high sensitivity and selectivity for TC, which will be used for the study of TC analysis in meat.