Blastocystis hominis provides major challenges for laboratory diagnosis due to its polymorphicnature in wet mounts which can result in confusion with other protozoa, yeast or even fatglobules. Studies revealed that simple smears were less sensitive than in vitro cultivation usingdifferent media for the detection of B. hominis in stool specimens. Cultures of B. hominis areusually enriched by different types of sera to enhance growth and multiplication of the parasite.The aim of this study is to assess the use of two sera types other than horse serum that iscommonly used in culture media for the growth, multiplication and detection of B. hominis inexamined stool samples and comparing the results with those obtained using horse serum.Fifty stool samples were collected from patients suffering from different colonic manifestationsattending Cairo University Hospitals. The samples were freshly cultured in three differentculture media using horse serum (in Jones’ medium), donkey serum (as a modification inJones’ medium) and human plasma (in modified Pavlova’s medium) in adequate preparations.Cultures were then left for incubation and examined by direct microscopy to detect Blastocystishominis.The results showed of 50 stool samples studied, 18 samples (36%) were positive results forB. hominis. The number of positive results obtained by horse serum, donkey serum and humanplasma were 13,18and 11 respectively. Paired comparisons were made between each 2 cultureswith each culture set as a reference once to detect the most appropriate one for diagnosis.When horse was set as the reference method, donkey serum showed a sensitivity of 100% andspecificity of 86.5% with a 90% agreement between the 2 methods. While human plasmashowed a sensitivity of 46.2% and specificity of 86.5% with an agreement of 76%. In addition,the vacuolar form was the commonest pattern observed in this study throughout all the three cultures.