An indirect competitive enzyme-linked aptamer assay（ic-ELAA） was developed for monitoring oxytetracycline（OTC） in complex food matrix.Various factors such as OTC-BSA concentration,aptamer concentration,avidin-HRP concentration,coating buffer,blocking agent,and binding buffer were evaluated for the performance of ELAA.The optimized ELAA showed a good sensitivity（IC50 of 6.3 ng/mL） and a wide linear range（0.5-50 ng/mL） for OTC.There was no obvious cross reactivity（CR） with most of the tetracyclines pesticides.Recoveries of OTC in food samples,including milk,milk powder,chicken,fish and honey,ranged from 62.1% to 102% with relative standard deviations（RSDs） less than 15%.In addition,the developed ELAA showed a good correlation with the high performance liquid chromatographic method（r2=0.979）.The results indicated that the developed method was a simple,sensitive,accurate and cost-effective approach for the screening of OTC in food samples.