The CellDirector 3D assay.

<p>a) Schematic overview of the CellDirector 3D assay setup for AST determination and b) close-up of the microfluidic growth chamber depicting details of the concentration gradient and MIC readout.</p

MIC values in mg/L obtained with the Etest and CellDirector 3D at bacterial inocula of 10⁵ CFU/mL.

<p>MIC values in mg/L obtained with the Etest and CellDirector 3D at bacterial inocula of 10⁵ CFU/mL.</p

Antibiotic concentration ranges used in CellDirector 3D.

<p>Antibiotic concentration ranges used in CellDirector 3D.</p

MIC over time for quality control strains.

<p>a) MIC determination and stabilisation over time for different inoculum sizes of <i>S</i>. <i>aureus</i> (VSSA) and <i>P</i>. <i>aeruginosa</i>, tested against vancomycin and ciprofloxacin, respectively. b) MIC over time for <i>E</i>. <i>coli</i> against ceftazidime, ciprofloxacin and tigecycline, <i>K</i>. <i>pneumoniae</i> against ciprofloxacin, and <i>P</i>. <i>aeruginosa</i> against ceftazidime, at an inoculum size of 10⁵ CFU/mL. Grey fields signify standard deviation (n = 3).</p

Vancomycin MIC values in mg/L as determined by the Etest, macrodilution and CellDirector 3D against VSSA and hVISA.

<p>Vancomycin MIC values in mg/L as determined by the Etest, macrodilution and CellDirector 3D against VSSA and hVISA.</p

Vancomycin MIC values (mg/L) for 13 <i>S</i>. <i>aureus</i> isolates from clinical blood cultures as determined by the Etest, macrodilution and CellDirector 3D (4 h readout).

<p>Vancomycin MIC values (mg/L) for 13 <i>S</i>. <i>aureus</i> isolates from clinical blood cultures as determined by the Etest, macrodilution and CellDirector 3D (4 h readout).</p

Growth rate analysis and MIC determination from CellDirector 3D images.

<p>Example of (a) a dark field image of the cell culture chamber after 300 min in the CellDirector 3D assay and (b) the corresponding plot of greyscale intensity change from the five images over the last 40 min (i.e. growth rate) for <i>S</i>. <i>aureus</i> (VSSA) and vancomycin. The vancomycin concentration on the x-axis of the plot corresponds to a pixel coordinate on the image x-axis. The MIC value (arrow) is set as the first position at which the change in greyscale intensity is below 0.1 (grey line). In the bottom panel (c) the computed MIC value is plotted for each time point. The MIC is not calculated (and is plotted as 0) until a growth rate signal above 0.1 is detected (at 160 min in the example above) and is stable after this point. The grey dotted line indicates the MIC at 5 h for this strain and antibiotic.</p