A total of 123 clinical Escherichia coli and Klebsiella spp. isolates were included in the study in order to evaluate VITEK 2 AST-NO29 (Nordic) card for detection of extended-spectrum beta-lactamases (ESBL) and to compare the results with genotypic ESBL verification. The results were also compared to alternative phenotypic methods, i.e. agar dilution and disk diffusion. The strains that were ESBL-positive according to AST-N029 were further analysed with the ESBL test card, VITEK 2 AST-N041. Using genotype as reference, Vitek 2 AES had the highest accuracy of the tested methods in classifying the strains as ESBL-positive or -negative (91.1%). When VITEK 2 gave ESBL as the only option for E. coli or K. pneumoniae, 44 of 45 (97.8%) strains had an ESBL gene. VITEK 2 achieved an accuracy of 94.9% and disk diffusion 95.9% compared to the agar dilution method as the phenotypic reference method for the E. coli and K. pneumoniae strains. For the K. oxytoca strains VITEK 2 achieved the highest accuracy (84.0%) of the methods used in this work.