Cell culture and FFA treatment in HepG2 cells using cultured cells were treated with various concentrations (0.05–0.5 mM) of long chain FFA with different degrees of saturation, including palmitate and stearate (saturated), palmitoleate and oleate (one double bond), and linoleic (two double bonds) (Sigma) in media containing 1% bovine serum albumin for times ranging up to 24 h (0.5, 1, 2, 4, 8, 16, and 24 h).
If you go to medline and search for Lee W-N.P. or Boros LG publications you will find many papers using HepG2 cells and stable isotope tracer based metabolomics (isobolomics) to measure fatty acid de novo synthesis, chain elongation and oxidation (beta carbons). You can also look for rosiglitazone treatment like in the paper I am attaching. LGB
salam Alaikum
Eungyeong Jang and research team about revealed Cell viability and lipid accumulation by MTT assay and Oil Red O staining method .
MTT assay method :
It was dissolved as a 1 mg/ml stock in phosphate-buffered saline (PBS). HepG2 cells were seeded at a density of 1.5 × 103 cells/well in 96-well plate and incubated for 48 h. The cells were treated with disparate concentrations of AC extract (100, 500 and 1000 μg/ml) for 24 h and FFAs 1 mM in another group for 1 h and 24 h, respectively. And then, 100 μl of MTT solution were treated for 2 h. After 4 h, MTT solution was removed by aspiration, the insoluble formazan crystals were dissolved in DMSO and measured the absorbance was read at 570 nm with a Bio-Rad model 680 microplate reader (Bio-Rad, Hercules, CA).
Oil Red O staining method :
Oil Red O solution was purchased from Sigma-aldrich. Oil Red O staining was performed according to the reference [24,25]. To stain adipocytes, cells were washed three times with PBS to remove unbound staining and fixed with 10% formalin for 1 h. After washing for three times with distilled water, cells were washed with 60% isopropanol briefly and incubated with 60% filtered Oil Red O solution (0.7 g per 200 ml of isopropanol) for 30 min at room temperature. Cells were washed with water briefly and then stained with hematoxylin (Sigma-Aldrich, USA). For quantitative analysis of Oil Red O contents levels, isopropanol was added to each samples and then shaken at room temperature for 5 min. The absorbance was read at 510 nm with a Bio-Rad model 680 microplate reader.
The article in attachments have all details
Good luck
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