The aim of this study was to evaluate a Chemiluminescence Enzyme Immunoassay (CLIA) developed for the detection of E. coli O157:H7, using different E. coli O157 serotypes. The sensitivity and specificity of the kit were determined from the tenfold dilutions of the 24-hour broth cultures of the test strains. According to the results obtained in this trial, the sensitivity of the kit is 103–104 cells ml−1, and it is specific for E. coli O157. Twenty-five g ground raw beef samples were prepared and inoculated with E. coli O157:H7 at different CFU g−1. The samples were incubated in 225 ml of modified E. coli broth with novobiocin (mEC + n) at 42 °C for 4 h and the immunoassays were performed following the instructions of the manufacturer. According to the results obtained by the CLIA test 101–102E. coli O157 g−1 can be detected from the sample. So this kit seems to be suitable for screening the samples before selective cultivation of E. coli O157:H7.