Fatty acid accumulation in RAW264.7 cells was measured by exposing the cells to oleic acid (OA), following the previously described method (19 (link)). RAW264.7 cells were exposed to 0.1 mM OA in the presence or absence of GlcN, galactosamine (GalN), mannosamine (ManN) or LPS for 24 hrs. Cells were subsequently washed and stained with 0.3% ORO working solution at RT for 15-30 min.
The rate of fatty acid uptake was determined using green fluorescent fatty acid, 4,4-difluoro-5,7-dimethyl-4-bora-3α,4α-diaza-s-indacene-3-hexadecanoic acid (BODIPYTM FL C16, Thermo Fisher Scientific, MA, USA). Cells were treated with LPS and/or GlcN for 12 hrs, washed twice with PBS, and incubated with 1 μg/ml BODIPYTM FL C16 for 3 min at RT. After an four washes, the fluorescence/well was measured (S1LFA; Biotek, VT, USA).
The rate of fatty acid uptake was determined using green fluorescent fatty acid, 4,4-difluoro-5,7-dimethyl-4-bora-3α,4α-diaza-s-indacene-3-hexadecanoic acid (BODIPYTM FL C16, Thermo Fisher Scientific, MA, USA). Cells were treated with LPS and/or GlcN for 12 hrs, washed twice with PBS, and incubated with 1 μg/ml BODIPYTM FL C16 for 3 min at RT. After an four washes, the fluorescence/well was measured (S1LFA; Biotek, VT, USA).
