Nerve Growth Factor (NGF) is the archetypal neurotrophin of a family of polypeptides .It has long occupied a critical role in developmental and adult neurobiology for its many important regulatory functions on the survival, growth and differentiation of nerve cells in the peripheral and central nervous system. The Caspase 3 (CASP3) is high affinity receptor for Nerve Growth Factor，thus a functional binding ELISA assay was conducted to detect the interaction of recombinant mouse NGF and recombinant human CASP3. Briefly, NGF was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to CASP3-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-NGF 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 mouse NGF and recombinant human CASP3 was shown in Figure 1, the EC50 for this effect is 0.86 ug/mL.