The demand for meat is expected to exceed production capacity by livestock in the coming decennia. Therefore, cultured beef might be a viable alternative to traditional livestock-derived beef. One of the problems however is the sustainability of cultured beef through the use of fetal bovine serum. We aimed to identify a serum-free medium or a serumreplacement that is as effective as the current method used for culturing bovine myoblasts. Cells were harvested from a female Blanc Bleu Belge cow and myoblasts were subsequently isolated. Cells were cultured in either Advanced DMEM containing 20% FBS and 10% HS or one of the chemically-defined, serum-free media for 6 days. MTS was used as a measure of cell proliferation at day 1, 4 or 6 and microscopic pictures were taken to assess cell morphology. FBM TM , TesR TM and Essential 8 TM are commercially available xeno-free media developed for human PSCs and fibroblasts, with the highest potential to sustain bovine myoblast proliferation. Of the supplements tested, XenoFree TM and a customprepared growth factor mix failed to stimulate cell proliferation. LipoGro TM stimulated cell proliferation in some cases but also changed the phenotype of myoblasts to an adipocyte-like phenotype. We conclude that serum-free media stimulate exponential cell expansion, albeit not to the extent of the current growth medium containing up to 30% serum. Further research is needed to investigate whether prolonged cell culture or an adaptation period could further increase cell proliferation.