Abstract
Most bacteria entering the bloodstream will be eliminated through complement activation on the bacterial surface and opsonophagocytosis. However, when these protective innate immune systems do not work optimally, or when bacteria are equipped with immune evasion mechanisms that prevent killing, this can lead to serious infections such as bacteremia and meningitis, which is associated with high morbidity and mortality. In order to study the complement evasion mechanisms of bacteria and the capacity of human blood to opsonize and kill bacteria, we developed a versatile whole blood killing assay wherein both phagocyte function and complement activity can easily be monitored and modulated. In this assay we use a selective thrombin inhibitor hirudin to fully preserve complement activity of whole blood. This assay allows controlled analysis of the requirements for active complement by replacing or heat-inactivating plasma, phagocyte function and bacterial immune evasion mechanisms that contribute to survival in human blood.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adult
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Biological Assay*
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Complement Activation*
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Complement C3 / chemistry
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Escherichia coli / growth & development
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Escherichia coli / immunology
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Female
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Fibrinolytic Agents / chemistry
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Fluoresceins / chemistry
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Fluorescent Dyes / chemistry
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Haemophilus influenzae / growth & development
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Haemophilus influenzae / immunology
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Hirudins / chemistry
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Humans
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Immune Evasion*
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Immunoglobulin G / chemistry
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Immunoglobulin M / chemistry
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Klebsiella pneumoniae / growth & development
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Klebsiella pneumoniae / immunology
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Leukocytes, Mononuclear / immunology*
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Leukocytes, Mononuclear / microbiology
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Male
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Neisseria meningitidis / growth & development
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Neisseria meningitidis / immunology
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Phagocytosis / immunology*
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Pseudomonas aeruginosa / growth & development
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Pseudomonas aeruginosa / immunology
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Staphylococcus aureus / growth & development
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Staphylococcus aureus / immunology
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Streptococcus pneumoniae / growth & development
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Streptococcus pneumoniae / immunology
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Succinimides / chemistry
Substances
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5-(6)-carboxyfluorescein diacetate succinimidyl ester
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Complement C3
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Fibrinolytic Agents
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Fluoresceins
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Fluorescent Dyes
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Hirudins
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Immunoglobulin G
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Immunoglobulin M
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Succinimides