Gentamicin protection assay

THP-1 cells or BMMs plated at 0.2×10⁶ cells per well of a 96-well plate were infected with L. monocytogenes or GFP-L. monocytogenes, or Francisella novicida at m.o.i. of 10 in 100 μl total volume for 1 h. Medium was replaced with 100 μg/ml of gentamicin-containing medium. After an additional 5 min (considered as 1-h time point) and 1, 2, 4, and 6 h (considered as 2-, 3-, 5-, and 7-h time point), cells were washed three times with 200 μl PBS and lysed in 100 μ l of 0.1% Triton X-100 in PBS. Tenfold serial dilutions of the lysates were prepared in PBS and 10 μl was dropped on a LB agar plate with no antibiotics. Colonies were counted after 36 h incubation at 37°C. Each experiment was performed in triplicate wells and repeated three times.