The monoclonal antibody RDIII-7 recognizes complement decay accelerating factor (DAF), also designated as CD55. RDIII-7 recognizes the common extracellular SCR region of CD55, detecting all isoforms of the protein. Cells express on their surface several proteins which protect against complement attack, namely C receptor I (CR1), decay accelerating factor (DAF), membrane cofactor protein (MCP) and CD59. CR1, DAF and MCP regulate the activation pathways of complement by either accelerating decay of the C3 and C5 convertase (CR1, DAF), or acting as cofactors for the serine protease factor I, which cleaves and irreversibly inactivates C3b (CR1, MCP).
Rat DAF (CD55) is a 60 kDa transmembrane protein that binds C3b and C4b to inhibit formation and half-life of the C3 convertases. It belongs to the receptors of complement activation (RCA) family. DAF is broadly distributed among cells in contact with plasma complment proteins, including both haematopoietic and nonhaematopoietic cells. Although DAF does not have an essential role in controlling hemolysis of erythrocytes, it has an important role in regulation of the deposition of C3 on nucleated cells.
Together with other complement regulators DAF protects self cells from autologous complement-mediated injury. DAF cooperates with CD46 in circumventing autologous C3 deposition, while CD59 inhibits the pathway at the critical end-point.
Flow cytometry, Frozen sections, Functional studies, Immuno fluorescence, Western blot
W: non-reduced with a band size of 60-70kDa. As positive control CHO-rat DAF hyper-expressing cells were used.
FS: RDIII-7 antibody completely blocked the protective effect of expressed rat DAF
For immunohistology, flow cytometry and Western blotting, dilutions to be used depend on detection system applied. It is recommended that users test the reagent and determine their own optimal dilutions. The typical starting working dilution is 1:50. For functional studies, in vitro dilutions have to be optimized in user's experimental setting.
As positive controle CHO-rat DAF hyperexpressing cells were used.