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March 09, 2017

Complement dependent cytotoxicity (CDC)

Background:

The aim of this assay is to test the complement activation ability of test antibodies or protein-based drugs.

After binding to the cell surface antigen IgG and IgM, the test item can activate the classical pathway of the complement system and thus leads to the formation of the membrane attack complex (MAC). By forming a pore in the membrane of the target cell, the lysis of the cell is induced.

Method:

The assay is carried out by co-incubating the target cell line in the presence of the antibody (at different concentrations) with normal human sera (NHS, the source of the active complement) or with heat inactivated NHS (iNHS, no active complement components). The percentage of cells killed can be determined by flow cytometry using a fluorescent dead-cell dye (7-AAD).

Outcome:

Cells are stained with the dead cell dye 7-AAD and samples are subjected to flow cytometry analysis. 7-AAD+ positive cell population represents the dead cells indicating the cytolytic effect of the complement.

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