Isolation and functional assay of the membrane complement inhibitors CD55 (DAF) and CD59 (MIRL).
The complement system is the primary effector of humoral immunity. Because of its enormous destructive capacity, mechanisms for confining the activity of the system to the desired target and elaborate safeguards for protecting self against complement-mediated injury have evolved. Human cells, particularly those found at sites of inflammation (e.g., hematopoietic and endothelial cells), express highly specialized membrane constituents that act independently or in concert with plasma regulatory proteins to inhibit the functional activity of complement. Decay-accelerating factor (DAF), or CD55, directly inhibits the formation and stability of the amplification C3 and C5 convertases of both the classical and the alternative pathways. Failure of a cell to regulate the amplification C3 and C5 convertases allows the generation of the potentially cytolytic membrane attack complex (MAC), or C5b-9 (consisting of the complement components C5b, C6, C7, C8, and C9). The primary cellular regulator of the MAC is the membrane inhibitor of reactive lysis (MIRL), or CD59, which restricts complement-mediated lysis by blocking assembly of the MAC (primarily at the stage of C9 binding and polymerization). This unit provides a basic protocol for isolating CD55 and CD59, along with two support protocols describing separate functional assays for CD59 and CD55.
Parker
Charles J
CJ
University of Utah School of Medicine and The Veterans Affairs Medical Center, Salt Lake City, Utah.
D018958:CD55 Antigens / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification*; D018957:CD59 Antigens / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification*; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D051916:Membrane Cofactor Protein / Q000737:chemistry / Q000276:immunology; D017463:Receptors, Complement 3b / Q000737:chemistry / Q000276:immunology; D012680:Sensitivity and Specificity
D018958:CD55 Antigens; D018957:CD59 Antigens; D051916:Membrane Cofactor Protein; D017463:Receptors, Complement 3b