Phosphatidylcholine assay kit

Cell-based fluorometric assays were performed using the Free Fatty Acid Uptake Assay kit (#ab176768, Abcam). Briefly, iHLCs were grown in 96 well plate and incubate 1 h in serum-free media. Fatty acid dye-loading solution was added for 4 h and the fluorescence signal measured with a SpectraMax fluorescence microplate reader at Ex/Em = 485/515 nm.
Triglyceride content of iHLCs was measured using the Triglyceride Quantification Colorimetric/Fluorometric kit (# K622-100, Biovision). iHLCs(10⁶ cells) were suspended and lysed in 5% NP-40/ddH2O solution. The output was measured at OD 570 nm for colorimetric assay after mixing and incubating with assay reagents provided with the kit.
Phosphatidylcholine in iHLC cell lysates was measured using the Phosphatidylcholine Assay kit (#ab83377, Abcam) and a SpectraMax Microplate reader (Molecular Devices). Briefly, iHLCs were grown in 96 well plates, washed in PBS and suspended in assay buffer provided with the kit. Both standards and samples were incubated with kit components as directed and the measurement was performed at OD570 (OD is optical density).

Phosphatidylcholine level in liver tissues was measured by colorimetric using phosphatidylcholine assay kit (ab83377, from Abcam, Cambridge, UK). Liver tissues were washed with cold PBS, resuspended in the assay buffer provided by the kit, and homogenized with a Dounce homogenizer on ice. Thereafter, samples were centrifuged for 5 min at 4°C at 12,000 × g to exclude the insoluble material and collect the supernatant. The supernatant was incubated on a 96-well plate with the reaction mix for 30 min at room temperature and were protected from light. The colorimetric reaction was measured at 570 nm. Optical density was measured using a microplate reader. Then the concentration of phosphatidylcholine was estimated.