FABP4 is a marker for adipocyte differentiation. It is induced during in vitro transdifferentiation of bovine pre-adipocytes to adipocytes by the presence of bovine serum lipids (BSL) in the absence of fetal bovine serum (FBS). The intention of this study was to test whether ascorbic acid (AA) has an influence on transdifferentiation and FABP4 expression. Subcutaneous adipose tissue was collected from calves. After induction of differentiation, adipocytes were incubated with or without AA (40 µL/mL), BSL (10 µL/mL) and/or FBS (10%) for 14d. The accumulation of non-polar lipids was evaluated by Nile-red fluorescence and normalized to DAPI fluorescence. Stem cell markers and FABP4 were investigated by quantitative RT-PCR and immunohistochemistry. Statistics was conducted by 2-way ANOVA. Results: The development of lipid droplets was promoted (P < 0.001) by BSL in absence of FBS. The mRNA expression of ENG was reduced in all treatments compared with pre-adipocyte values, with lowest values in FBS-free media (P < 0.05). The mRNA expression of NT5E was not reduced in cells incubated with FBS compared with pre-adipocyte values but was reduced in FBS-free media compared with pre-adipocyte values and most FBS-treated groups (P < 0.05). In the presence of AA, the mRNA expression of the adipocyte marker FABP4 increased 208, 30 and 144 times in media containing BSL, FBS and BSL+FBS, respectively (P < 0.05). In the absence of AA, only BSL increased the mRNA expression of FABP4 (75 times) above pre-adipocyte values (P < 0.05). The differential mRNA expression of FABP4 was mirrored by immunohistochemical staining of the FABP4 protein, which was most intense in cultures supplemented with BSL and AA in the absence of FBS. Conclusions: The study confirmed that FBS inhibits bovine adipocyte differentiation whereas BSL potently induces such differentiation. A combination of BSL and AA was most effective in inducing the adipocyte marker FABP4 despite the fact that lipid accumulation was not significantly influenced by the co-presence of AA. This work was supported by the Elsa-Neumann-Stipendium.