Active Major Basic Protein (MBP)

Myelin basic protein (MBP), a 18.5-kDa encephalitogenic peptide, is the second most abundant in central nervous system and responsible for adhesion of the cytosolic surfaces of multilayered compact myelin, MBP is biased expressed in cerebellum adult and cortex adult. It can interact with a number of polyanionic proteins including actin, tubulin, Ca(2 )-calmodulin, and clathrin, and negatively charged lipids, and acquires structure on binding to them. Calmodulin Like Protein 3 (CALML3) is a kind of Ca(2 )-calmodulin, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human MBP and recombinant human CALML3.Briefly, MBP was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to CALML3-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-MBP 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 450/630nm immediately. The binding activity of ecombinant human MBP and recombinant human CALML3 was shown in Figure 1, the EC50 for this effect is 0.081µg/mL..