Background: Escherichia coli (E. coli) is a Gram-negative bacterium that is facultative anaerobic in nature. E. coli strains causing urinary tract infection (UTI) are called uropathogenic E. coli (UPEC). UPEC makes up to 70-95% of UTIs. UTI is a serious health problem concerning antibiotic resistance and biofilms formation. Biofilm can restrict the diffusion of antimicrobials. Aim: To detect E. coli biofilm producers to improve prognosis and treatment and reduce morbidity and mortality rates due to UTI. Methods: Forty-seven clinical isolates of non-duplicated UPEC were collected from clinically suspected cases of UTI in Suez Canal University Hospitals (SCUHs). Isolates were identified as E. coli by colony morphology, Gram staining and biochemical reactions. Antimicrobial susceptibility testing was performed by Kirby–Bauer disc diffusion method on Muller–Hinton agar plate. The detection of biofilm was carried out by Congo red agar (CRA) method and modified tissue culture plate (MTCP) method. Results: The CRA method detected 63.8% of E. coli isolates as biofilm producers while the MTCP method detected 40.4% of them as biofilm producers. The sensitivity, specificity and accuracy of CRA method in comparison to MTCP method were 100%, 60.7% and 76.6% respectively. All biofilm producers and non-producers were resistant to amoxicillin-clavulanate and ceftazidime. The maximum sensitivity was seen for imipenem (91.49 %), followed by nitrofurantoin (78.72%) and then gentamicin (63.83%). Biofilm producers showed significant resistance (73.68%) to norfloxacin when compared to nonbiofilm producers (25%) (P=0.003). Conclusion: The prevalence of biofilm production among UPEC in SCUHs is 40.4%. Imipenem is the drug of choice for treating biofilm producing UPEC.