Aims We developed a new method for detection of the intracellular parasite, Nosema ceranae, one of the most economically devastating pathogens of the honeybee. Methods and Results The SWP-32 antibody was used for the development of an enzyme-linked immunosorbent assay (ELISA). We also compared the efficiency of this ELISA to microscopy and quantitative real-time (qRT) PCR, the methods currently in use. Conclusions ELISA is comparable in sensitivity with the qRT-PCR, less expensive and faster. When this method is commercialized and made available to bee-keepers, it will allow them to make informed decisions for the application of in-hive chemicals. Hence, bee-keepers may be able to determine when treatments for control of N.ceranae are unnecessary and reduce the cost, time and possible side effects of these treatments. Significance and Impact of the Study This assay provides the first serological method for detection of N.ceranae in bee colonies, which is as sensitive as DNA amplification. It can be easily adopted for both laboratory and field applications.