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A novel method for direct measurement of complement convertases activity in human serum

Lookup NU author(s): Professor Tim Goodship, Professor Kevin MarchbankORCiD

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Abstract

Complement convertases are enzymatic complexes that play a central role in sustaining and amplification of the complement cascade. Impairment of complement function leads directly or indirectly to pathological conditions, including higher infection rate, kidney diseases, autoimmune-or neurodegenerative diseases and ischaemia-reperfusion injury. An assay for direct measurement of activity of the convertases in patient sera is not available. Existing assays testing convertase function are based on purified complement components and, thus, convertase formation occurs under non-physiological conditions. We designed a new assay, in which C5 blocking compounds enabled separation of the complement cascade into two phases: the first ending at the stage of C5 convertases and the second ending with membrane attack complex formation. The use of rabbit erythrocytes or antibody-sensitized sheep erythrocytes as the platforms for convertase formation enabled easy readout based on measurement of haemolysis. Thus, properties of patient sera could be studied directly regarding convertase activity and membrane attack complex formation. Another advantage of this assay was the possibility to screen for host factors such as C3 nephritic factor and other anti-complement autoantibodies, or gain-of-function mutations, which prolong the half-life of complement convertases. Herein, we present proof of concept, detailed description and validation of this novel assay.

Publication metadata

Author(s): Blom AM, Volokhina EB, Fransson V, Stromberg P, Berghard L, Viktorelius M, Mollnes TE, Lopez-Trascasa M, van den Heuvel LP, Goodship TH, Marchbank KJ, Okroj M

Publication type: Article

Publication status: Published

Journal: Clinical & Experimental Immunology

Year: 2014

Volume: 178

Issue: 1

Pages: 142-153

Print publication date: 01/10/2014

Online publication date: 04/09/2014

Acceptance date: 20/05/2014

ISSN (print): 0009-9104

ISSN (electronic): 1365-2249

Publisher: Wiley-Blackwell Publishing Ltd.

URL: http://dx.doi.org/10.1111/cei.12388

DOI: 10.1111/cei.12388

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