Triglycerides and cholesterol kits

Chicken blood samples were collected from brachial wing veins using 23-gauge needles (Kelly and Alworth, 2013 (link)). The blood samples were transferred into EDTA-coated blood collection tubes (Becton Dickinson, Franklin Lakes, NJ) and mixed gently for 30 min. After centrifugation at 1,000 × g for 10 min, the supernatant fraction was collected for TG and cholesterol measurement using Triglycerides and Cholesterol kits (Randox Laboratories, County Antrim, UK). The detection kits were used according to the manufacturer's instructions. The signals were detected by an Epoch Microplate Spectrophotometer (BioTek, Winooski, VT) by measuring absorbance at 500-nm wavelength.

Twenty milligrams of chicken liver was homogenized in 1 mL of PBS. Five milliliters of chloroform/methanol (2/1, v/v) was added; samples were vortexed for 1 min followed by incubation at 4°C for 2 h. After centrifugation at 1,650 × g for 10 min the bottom phase was collected and dried using nitrogen gas. Isopropanol/Nonidet P-40 (9:1, v/v) solution was used to dissolve the lipid samples. The levels of liver TG and cholesterol were measured using Triglycerides and Cholesterol kits (Randox Laboratories). Values of liver TG and cholesterol were normalized with tissue weight.