Active Complement Component 8b (C8b)

Complement Component 8b (C8b), one of the three subunits of the complement component 8 (C8) protein, has only been identified in vertebrates. C8 is one of the five components (C5b, C6, C7, C8 and C9) that interact to form the MAC that plays a key role in the innate and adaptive immune response by forming pores in the plasma membrane of target cells. Besides, C9 can interact with C8b, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human C8b and recombinant human C9. Briefly, C8b was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to C9-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-C8b 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 C8b and recombinant human C9 was shown in Figure 1, the EC50 for this effect is 0.59 ug/mL.