Cholesterol absorption assay for Caco-2 cells

Caco-2 cells were grown in 48-well plastic dishes containing 0.5 mL of fetal bovine serum supplemented with DMEM for 14 days post-confluence. [¹⁴C]-Labeled micellar cholesterol uptake in Caco-2 cells for 120 min was measured as described previously¹¹. The [¹⁴C]-labeled micellar solutions (0.5 mL) contained casein tryptic hydrolysate (CTH), HPHU or FP, 5 g/L respectively. The [4-¹⁴C]-labeled micellar solution (6 mL) was prepared at the following concentrations and mixed by sonication (Taitec, VP-5): 3.7 kBq [4-¹⁴C]-cholesterol (2.0 Gbq/mmol, NEN), 2 µmol/L cholesterol (Sigma Aldrich), 6.6 mmol/L sodium taurocholate (Sigma Aldrich), 20 µmol/L oleic acid (Sigma Aldrich), 0.6 mmol/L phosphatidylcholine (Sigma Aldrich), 5 µmol/L monoolein (Sigma Aldrich). The cellular protein was determined using a commercially available kit (Bio-Rad Protein Assay; Bio-Rad).