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A high-throughput, bead-based, antigen-specific assay to assess the ability of antibodies to induce complement activation.

Journal of immunological methods (2019-07-14)
Stephanie Fischinger, Jonathan K Fallon, Ashlin R Michell, Thomas Broge, Todd J Suscovich, Hendrik Streeck, Galit Alter
ABSTRACT

The complement system plays a critical role in innate immune defense against pathogens, both via non-specific direct pathogen recognition and killing or via antigen-specific indirect recruitment by complement fixing antibodies. While various assays for measuring complement activation have been developed, few provide a high-throughput, sample-sparing approach to interrogate the qualitative differences in the ability of antibodies to drive complement activation. Here we present a high-throughput, sample-sparing, bead-based assay to evaluate antigen-specific antibody-dependent complement activation against nearly any antigen. Optimization of buffer composition, kinetics of immune complex formation, as well as complement source all contribute critically to the development of a robust, highly flexible and high-throughput approach to analyze antibody-dependent complement deposition (ADCD). Thus, the optimized bead-based, antigen-specific assay represents a simple, highly adaptable platform to profile antibody-dependent complement activation across pathogens and diseases.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
RPMI-1640 Medium, With sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
IgG from human serum, reagent grade, ≥95% (SDS-PAGE), essentially salt-free, lyophilized powder