Quantification of complement C5b-9 binding to cells by flow cytometry.

Area of Science:

• Immunology
• Cell Biology
• Molecular Biology

Background:

• The complement system is a key part of innate immunity, mediating host defense against pathogens and clearing damaged cells.
• Complement activation leads to the formation of the membrane attack complex (MAC), C5b-9, which can cause cell lysis at high concentrations or signal transduction at low concentrations.
• Understanding C5b-9 deposition is vital for assessing complement-mediated cellular responses and developing targeted therapies.

Purpose of the Study:

• To develop and detail a reliable method for quantifying C5b-9 complex deposition on cell surfaces.
• To enable accurate assessment of the complement-activating capacity of various cells and antibodies.
• To provide a protocol for researchers studying complement system interactions with cells.

Main Methods:

• Utilized flow cytometry for quantitative analysis of C5b-9 deposition.
• Employed a novel antibody specifically recognizing an antigen of the C5b-9 complex.
• Described a detailed experimental protocol for sample preparation and data acquisition.

Main Results:

• Successfully quantified the amount of C5b-9 complexes deposited on cell surfaces.
• Demonstrated the utility of the novel antibody for sensitive and specific C5b-9 detection.
• Established a robust flow cytometry-based assay for complement activation studies.

Conclusions:

• The developed flow cytometry protocol enables precise quantification of C5b-9 deposition.
• This method is valuable for evaluating cellular and antibody-driven complement activation.
• Accurate C5b-9 quantification aids in understanding complement's role in health and disease.