Lysophosphatidic acid phosphatase type 6 (ACP6), is a 44-47 kDa monomeric member of the histidine acid phosphatase family of proteins. ACP6 is highly expressed in kidney, heart, small intestine, muscle, liver, prostate, testis, ovary and exists as two isoforms as a result of alternative splicing events. It plays a role in regulating lipid metabolism that hydrolyzes LPA to monoacylglycerol (MAG) and phosphate. Annexin V (ANXA5) binds to phospholipid?bilayers, forming two-dimensional crystals that block the?phospholipids?from availability for coagulation enzyme reactions. A functional binding ELISA assay was conducted to detect the interaction of recombinant human ACP6 and recombinant mouse ANXA5. Briefly, ACP6 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to ANXA5-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-ACP6 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 ACP6 and recombinant mouse ANXA5 was shown in Figure 1, the EC50?for this effect is 0.08 ug/mL.