Introduction: Urinary tract infection (UTI) is one of the leading causes of morbidity encountered in clinical practice. Emerging resistance of the uropathogens to the antimicrobial agents due to biofilm formation is a matter of concern while treating symptomatic UTI which leads to longer stay in hospital and increased cost of treatment. Detection of biofilm producer strains will guide the clinician in modifying antibiotic therapy for better clinical management and also help in designing adequate control measures as the isolates are also resistant to variety of disinfectants. Aim: The present study was conducted to compare three methods Tissue Culture Plate (TCP) method, Tube method (TM) and Congo red agar (CRA) to detect biofilm formation by uropathogens and to correlate their susceptibility pattern with biofilm formation. Materials and methods: A total of 150 culture positive significant isolates from urine samples were subjected to biofilm detection by TCP method, Tube method and CRA method. TCP was considered as gold standard method. Impact of biofilm production was correlated with the antibiotic resistant pattern. Results: Out of 150 culture positive significant bacterial isolates, Gram negative organisms were isolated from 122 (81.33%) specimens and Gram positive growth was seen in 28 (18.67%) samples. Escherichia coli was the commonest Gram negative organism isolated (42.66%) while among Gram positive isolates, maximum biofilm production was shown by Enterococcus faecalis (66.66%). The gold standard TCP method detected 51 (34%) isolates as strong and 11(7.33%) isolates as moderate biofilm producers and remaining 88 (58.67%) isolates were weak/non-biofilm producing bacteria.