The monoclonal antibody 3/11 recognizes C3a, an activated fragment of the complement protein C3, a 190 kDa protein which is the most abundant complement protein in serum. C3 is the third component of the complement system and plays a central role in the complement cascade. The three pathways of compliment activation come together- at this key molecule. Cleavage of the pivotal complement component C3 results in the generation of a set of effector molecules which have diverse biological functions. The cleavage product C3b promotes complement acrtivation and the subsequent formation of the membrane attack complex. C3a, the small (ca. 10 kDa) cleavage product possesses anaphylatoxic as well as immunoregulatory- properties. It exerts its function through a specific receptor (C3aR), which belongs to the G-protein coupled receptor family. Expression of C3aR- has been reported in many cell types including myeloid and non-myeloid cells. Expression of C3aR on haematopoietic stem/progenitor cells has been shown to promote the trafficking/homing of these cells to the BM. Engagement of C3aR on DCs and T cells has been shown to up-regulate these cell functions.
The monoclonal antibody 3/11 reacts specifically with the C3a anaphylatoxin, providing a tool for monitoring acute complement activation (C3a generation) in experimental mouse models.
Flow cytometry, Functional studies, Immuno assays, Western blot
IA: Plates were coated with goat anti rat IgG-Fc antibody, then incubated with 1:100 hybridoma supernatant to form capture complex for C3a.
FS: Antibody 3/11 functions as an antagonist. The antibody can be used to block C3a (Ref.4).
W: A non-reduced sample treatment and 7.5% polyacrylamide gel was used. Proteins were blotted on nitrocellulose, blocked with 5% skimmed milk and incubated with 1:10 culturesupernatant for 1h at RT. The band size of the entire C3 molecule is 190 kDa (Ref.1).
For Western blotting (non-reduced) 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.