Objective To detect ESBLs in E.coli and K.pneumoniae with appropriate method.Methods The suspiciously produced ESBLs were detected by standard screen test among 110 E.coli and 84 K.pneumoniae. ESBLs possessing strains were confirmed by E test, single disc diffusion test with clavulanic acid or sulbatan, and two kinds of double disc synergy test. Results There was a significant difference between the detected rate of ESBLs in E.coli (16/26) and K. pneumoniae (4/20) by E test( P 0.05). The single disc diffusion test with clavulanic acid or sulbatan was applicable to confirm ESBLs in K. pneumoniae (12/20). Conclusions Single disc diffusion test is a sensitive, convenient and inexpensive method to confirm ESBLs in E.coli and K.pneumoniae in clinical laboratory.