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Cultivation of bloodstream forms of Trypanosoma brucei and T. evansi in a serum‐free medium

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Abstract:

Bloodstream forms of Trypanosoma brucei and T. evansi have been cultivated in an axenic culture system by using Iscove's modified DMEM‐based HMI‐9 medium supplemented with bathocuproinedisulphonic acid,l‐cysteine, hypoxanthine, 2‐mercaptoethanol, pyruvate, thymidine and 10% fetal bovine or adult horse serum. We developed a serum‐free medium (HMI‐244) in which serum in HMI‐9 was replaced by fatty acid‐free bovine serum albumin, bovine α2‐macroglobulin, bovine ‐lipoprotein, d‐biotin, retinol, ‐alanine, l‐anserine nitrate salt, l‐ornithine hydrochloride, O‐phosphorylethanolamine, sarcosine, taurine, adenosine‐ 5′‐triphosphate, 2′‐deoxycytidine‐5′‐monophosphate, 2′‐deoxyguanosine‐5′‐monophosphate, and 5‐methyltetrahydrofolic acid. Maximum cell densities and population doubling times were 2.6×106 cells/ml; 10.6 hours and 2.2×106 cells/ml; 10.9 hours for T. brucei and T. evansi, respectively. Bloodstream forms continued to proliferate in the serum‐free cultures for more than 90 days and the trypomastigotes retained their morphological characteristics and infectivity to mice. If validated, this serum‐free medium may help reduce future interlaboratory variability in biochemical, immunological, molecular biological and drug sensitivity studies on these parasites.

Keywords: T. brucei; T. evansi; cultivation; serum‐free medium

Document Type: Research Article

DOI: http://dx.doi.org/10.1046/j.1365-3156.1997.d01-268.x

Affiliations: 1: The Research Center for Protozoan Molecular Immunology, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido, Japan, 2: Walter Reed Army Institute of Research, Washington, District of Columbia, USA, 3: Department of Protozoology, The Institute of Tropical Medicine, Nagasaki University, Nagasaki, Japan, 4: Department of Veterinary Physiology, Obihiro University of Agriculture and Veterinary Medicine Obihiro, Hokkaido, Japan

Publication date: March 1, 1997

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