Fatty degeneration in skeletal muscle is a hallmark of many myopathies, sarcopenia, obesity and type-2 diabetes. It has been shown that human skeletal muscle fibroblasts (but not the myogenic cells) have the potential for transdifferentiation into adipocytes in culture, suggesting that these cells may be the cause of adipocyte accumulation in muscle (1). In order to study the transdifferentiation potential of these cells in vivo (xenotransplantation), it is important to genetically label the fibroblasts before transplanting them into the host species and to confirm that they still retain their potential for adipogenic transdifferentiation. In this study we have sought to transduce human primary skeletal muscle fibroblasts with a GFP lentivirus and determine if the transdifferentiation potential of human muscle-derived primary fibroblasts is retained. Following local anaesthesia (2% lidocaine), a muscle biopsy sample was obtained from the vastus lateralis muscle of a healthy, young, female subject (aged 20 years). Following isolation and expansion, cells were purified by immuno-magnetic cell-sorting using CD56 microbeads (2). The CD56-negative fraction (enriched for fibroblasts) was subsequently grown in skeletal muscle growth medium (PromoCell) and transduced with a GFP lentivirus at different doses (1:15, 1:30, 1:40 and 1:100). The transduction efficiency was measured by flow cytometry. For transdifferentiation, GFP-positive fibroblasts were exposed to 300 μM oleic acid and 300 μM palmitic acid complexed to BSA at 15 mg/ml in proliferation medium (1). The transduction efficiency was measured by flow cytometry 24 hours post-transduction for each GFP-lentiviral dose. The transduction efficiency for each dose was 72% (1:15), 60% (1:30), 50% (1:40) and 30% (1:100). The 1:15 dose appeared to compromise fibroblast viability, so the 1:30 dose was selected for further transductions. Using this dose, the transduction efficiency was 75% five days after transduction. Following treatment with fatty acids for 72 hours, cells were fixed and analysed by immunohistochemistry using antibodies against the adipogenic transcription factors C/EBPα and PPARγ, and Oil Red O which stains lipids. The results showed that applying fatty acids to GFP-positive fibroblasts resulted in their transdifferentiation into adipocytes as evidenced by a clear accumulation of Oil Red O positive lipid droplets and increased expression of the adipogenic transcription factors C/EBPα and PPARγ compared to the non-fatty acid treated GFP-positive fibroblasts. This study confirms that GFP lentivirus transduction of human primary skeletal muscle derived fibroblasts does not affect their transdifferentiation potential and can therefore be used to successfully label cells for future in vivo experiments.