Pentra 400

Cells were scraped and collected in PBS (200 µL per well in a 6-well plate format), and were centrifuged at 12500 rpm at 4°C for 10 min and the buffer was discarded by pipetting. Triglycerides were extracted by incubating samples with 200 µL of cold 100% isopropanol at 4°C for 2 h. After a centrifugation for 5 min at 1300 x g at 4°C, 250 µL of supernatant was used to analyze triglycerides on ABX Pentra 400 according to the method for triglyceride quantification in the liver. Another plate of cells (cultured in same conditions) were stained with DAPI and differences in cell number were corrected using DAPI fluorescence (measured at excitation 350 nm and at emission 450/490 nm). Images were taken using a bright field microscope (Zeiss Axio Vert).

Plasma concentration of BUN, triglyceride, and glucose were detected by a Fuji DriChem 4000i clinical chemistry analyzer (Scil, Viernheim, Germany). Plasma concentration of lactate, creatinine, and ALT were measured by an enzymatic-spectrophotometric assay using an ABX Pentra 400 instrument.

Linearity was evaluated by manual stepwise dilution of two canine lithium heparin plasma samples with markedly increased CRP concentration of ~ 245 and 360 mg/l determined on the ABX Pentra 400 analyzer which was used as a reference method. Serial dilution resulted in specimens with 1.0, 0.8, 0.6, 0.4, 0.2, 0.1, 0.05, and 0.025 of the original CRP concentration. All diluted aliquots were analyzed in triplicates in a single run. For all dilution steps % recovery rate was evaluated by comparison of expected and measured results. Furthermore, the dilution series was used for calculation of the recovery rate. The experiment was conducted twice to verify the measurements.

Blood samples were analyzed for FBS level and lipid profileS (TC, HDL-c, LDL-c and TG)) using ABX Pentra 400 machine. Enzymatic colorimetric assay method was used for the measurement of total cholesterol (CHODPAP method) and triglyceride (GPO-PAP method), while direct homogeneous enzymatic colorimetric assay technique was utilized for the measurement of HDL-c and LDLc. FBG level was measured by glucose oxidase method (GOD-PAP).

A trained technician followed stringent standard operating procedures and collect 5 milliliters of venous blood specimen from each study participant after they had fasted for the previous night. The sample was then stored at room temperature for 30 min before being centrifuged using a Rotanta 960 centrifuge for 5 min at a speed of 4000 revolutions per minute. By using the direct end- point enzymatic approach, the ABX Pentra 400 automated clinical chemistry analyzer (Spain) examined the serum lipid profiles (TC, HDL-c, LDL-c, and TGs).