Biofilm is one of the known virulence factors of Candida, an important pathogen and commensal. Early detection of biofilm production may be useful for clinical decision because of its suggestive property for potential pathogenic capacity of Candida isolates. In this study, we simultaneously screened 411 clinical isolates of Candida spp. by microtiter plate (MTP), visual tube (TM) and congo red agar (CRA) methods for determining their ability to form biofilm and also evaluated the reliability of these methods in order to determine most suitable screening method. The accuracy of TM and CRA methods were also evaluated by using MTP as reference method. Of the 411 Candida spp., 159 (38.7%) displayed a biofilm-positive phenotype by MTP method. Sensitivity and specificity of the tube method were 68 and 98%, respectively and the positive predictive value (PPV) was 97% and the negative predictive value (NPV) was 83%. The sensitivity for congo red agar test was 79% and the specificity was 99%, and the PPV was 99% and the NPV was 88%. Tube method showed very good agreement for the isolates producing strong biofilm, whereas differentation of isolates producing weak biofilm was difficult. By the congo red method, classification of existing biofilm was problematic. Among the three methods studied, microtiter plate method may be suggested as the most sensitive method, which is easy to conduct and applicable as a routine process.