Bacteria were grown to mid-log phase in Frantz media supplemented with 4 mM D,L-lactate (Sigma-Aldrich) and 2 mM cytidine 5′-monophospho-N-acetyl-neuraminic acid (CMP-NANA; Carbosynth) to enhance sialylation of lipooligosaccharide (55 (link)). Test sera were heated for 30 minutes at 56°C to inactivate intrinsic complement. The exogenous source of human complement was human serum depleted of IgG with a protein G column (HiTrap Protein G HP, 5 ml; GE Healthcare) (12 (link)). Serum bactericidal antibody titers were assigned as the interpolated dilution resulting in 50% survival of the bacteria, compared with CFU/ml of bacteria incubated at 37°C for 60 minutes with negative control sera and complement.
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Serum Bactericidal Antibody Titers
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Other organizations : UCSF Benioff Children's Hospital
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Variable analysis
independent variables
- Bacteria grown to mid-log phase in Frantz media
- Supplementation of Frantz media with 4 mM D,L-lactate (Sigma-Aldrich)
- Supplementation of Frantz media with 2 mM cytidine 5′-monophospho-N-acetyl-neuraminic acid (CMP-NANA; Carbosynth)
- Serum bactericidal antibody titers
- Heating of test sera for 30 minutes at 56°C to inactivate intrinsic complement
- Human serum depleted of IgG using a protein G column (HiTrap Protein G HP, 5 ml; GE Healthcare) as the exogenous source of human complement
- Bacteria incubated at 37°C for 60 minutes with negative control sera and complement
- Bacteria incubated at 37°C for 60 minutes with negative control sera and complement
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