CULTIVATION OF BLOOD-STREAM FORMS OF TRYPANOSOMA-BRUCEI AND T-EVANSI IN A SERUM-FREE MEDIUM

Bloodstream forms of Trypanosoma brucei and T. evansi have been cultiv ated in an axenic culture system by using Iscove's modified DMEM-based HMI-9 medium supplemented with bathocuproinedisulphonic acid, L-cyste ine, hypoxanthine, 2-mercaptoethanol, pyruvate, thymidine and 10% feta l bovine or adult horse serum. We developed a serum-free medium (HMI-2 44) in which serum in HMI-g was replaced by fatty acid-free bovine ser um albumin, bovine alpha(2)-macroglobulin, bovine beta-lipoprotein, d- biotin, retinol, beta-alanine, L-anserine nitrate salt, L-ornithine hy drochloride, O-phosphorylethanolamine, sarcosine, taurine, adenosine- 5'-triphosphate, 2'-deoxycytidine-5'-monophosphate, 2'-deoxyguanosine- 5'-monophosphate, and 5-methyltetrahydrofolic acid. Maximum cell densi ties and population doubling times were 2.6 X 10(6) cells/ml; 10.6 hou rs and 2.2 x 10(6) cells/ml; 10.9 hours for T. brucei and T. evansi, r espectively. Bloodstream forms continued to proliferate in the serum-f ree cultures for more than go days and the trypomastigotes retained th eir morphological characteristics and infectivity to mice. If validate d, this serum-free medium may help reduce future interlaboratory varia bility in biochemical, immunological, molecular biological and drug se nsitivity studies on these parasites.