Background Endophthalmitis is, although relatively rare, a serious intraocular infection, which could result in a loss of visual function. Therefore, the rapid diagnosis and initiation of the appropriate treatment is of critical importance. To date, approximately 60 percent of eyes with a clinical diagnosis of infectious endophthalmitis show a posit ive microscopic or culture result. By using the very sensitive polymerase chain reaction this number might increase. Patients and methods In a series of 12 eyes with infectious endophtalmitis we have performed microscopic investigations, diagnostic culture iud polymerase chain reaction in aqueous humor and vitreous in order to detect the infectious agent. Results Microscopic investigations showed a positive result in the vitreous of 3 eyes. This number improved to 6 eyes using culture media. Significant less positive results were obtained in the aquous humor, The infectious agent could be detected in the aquous humor in all 12 eyes and in the vitreous in 9 eyes by PCR. Only in 2 eyes with a delayed endophthalmitis the vitreous was negative. Conclusions The detection of the infectious agents was more, successful using PCR compared to conventional microbiological tests. In particular, for the diagnosis of delayed endophthalmitis PCR proves to be very superior, in all cases of delayed endophthalmitis the pathogen could be detected in the aquous humor.