Twenty-six nasal and oropharyngeal swabs were collected from fatty buffalo calves in a herd at Ismailia Governorate suffering from pneumoenteritis with no response to antibiotic treatment and 40 milk samples collected from buffalo dairy herd suffering from mastitis at Beni suef Governorate, Egypt. The samples were submitted to laboratory diagnosis for the detection of mycoplasma infection. Using the culture method, 13 oropharyngeal and nasal swabs were positive, 4 were antigenically related to Mycoplasma bovirhinis, 2 were M. bovigenitalium, 2 were A. modicum and 5 were A. laidlawii Regarding the milk samples, 10 were positive; 2 were M. bovirhinis,5 were M. bovigenitalium and 3 were M. bovis. Examination of these samples with immunobinding assay (IBA) gave the same number of positive samples and the same species of mycoplasma. The IBA proved to be as sensitive and as specific as the culture. method. Moreover, IBA is more simple and have the advantage of utility and rapidity, 2 hours are needed to complete diagnosis instead of 10 days in case of the culture method.