Active Apolipoprotein A5 (APOA5)

Apolipoprotein A5 (APOA5) is a small protein, expressed predominantly in the liver. It has been identified to play an important role in lipid metabolism, specifically in triglyceride (TG) and TG-rich lipoproteins (TRLs) metabolism. Apolipoprotein C3 (APOC3) has been proved to be one of the ligands of APOA5. Thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human APOA5 and recombinant human APOC3. Briefly, APOA5 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to APOC3-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-APOA5 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µL stop solution to the wells and read at 450/630 nm immediately. The binding activity of recombinant human APOA5 and recombinant human APOC3 was shown in Figure 1, the EC50 for this effect is 0.14 ug/mL.