Bacterial killing assays in ecoimmunology require cross-validation by agreement statistics

The bacterial killing assay (BKA) is a simple assay that assesses the
ability of the constitutive immune system, typically the plasma-based
immune system, to clear pathogens in vitro. Historically, this has been
accomplished through counting bacterial colonies on solid agar plates
(CFU-based), however, over the last 15 years, a technique that makes use
of optical density (OD-based) has been steadily gaining in popularity.
Although cross-validations have been performed between these two methods
in the past, they only compared measurements of growth (colony counts and
absorbance) using correlation statistics with plasma. We performed our own
method cross-validation using first blood cells (red and white blood cells
with plasma removed) and then plasma, and compared measurements of growth
and bactericidal ability using both correlation and agreement statistics.
Our results found that while there was a weak correlation between colony
counts and absorbance at 300nm, we found poor agreement between techniques
in the cell-based BKA, with slightly stronger agreement when done with
plasma. We believe this is caused by immunological processes, especially
in the case of the cell-based BKA, changing the optical properties of the
culture.