Antimicrobial Susceptibility Assay

S. aureus strains carrying the pSB2019:tcaA plasmid were grown overnight in TSB with 10ug/ml chloramphenicol. Each strain was then normalized to an OD_600nm of 0.05 in fresh TSB and subcultured to an OD_600nm of 0.5–0.6. Cultures were washed in PBS and concentrated to an OD_600nm of 1 in PBS. 100 ul of bacteria was combined with 100 ul of the appropriate antimicrobial compound (arachidonic acid or LL-37) in a black 96-well plate. GFP Fluorescence (485_nm excitation/520_nm emission/1000 gain) was measured in a PHERAstar FSX plate reader (BMG Labtech) over 2.5 h (readings every 30 min with 200 rpm shaking).

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Douglas E.J., Palk N., Brignoli T., Altwiley D., Boura M., Laabei M., Recker M., Cheung G.Y., Liu R., Hsieh R.C., Otto M., O’Brien E., McLoughlin R.M, & Massey R.C. (2023). Extensive re-modelling of the cell wall during the development of Staphylococcus aureus bacteraemia. bioRxiv.

Publication Preprint 2023

PHERAstar FSX

Antimicrobial Arachidonic acid Bacteria Chloramphenicol Fluorescence Plasmid S aureus Strain

Corresponding Organization : University College Cork

Other organizations : University of Bath, University of Milan, University of Exeter, University of Tübingen, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Trinity College Dublin

Top 5 similar protocols

Variable analysis

independent variables

Antimicrobial compound (arachidonic acid or LL-37)

dependent variables

GFP Fluorescence (485nm excitation/520nm emission/1000 gain)

control variables

S. aureus strains carrying the pSB2019:tcaA plasmid
Overnight growth in TSB with 10ug/ml chloramphenicol
Normalization to OD600nm of 0.05 in fresh TSB
Subculture to OD600nm of 0.5-0.6
Washing in PBS
Concentration to OD600nm of 1 in PBS
Measurement in a black 96-well plate
Measurement in a PHERAstar FSX plate reader (BMG Labtech) over 2.5 h (readings every 30 min with 200 rpm shaking)

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