Normalizing spot intensity by spot area increases precision of growth curve analysis. An overnight culture was diluted 1:2000 and distributed into a 96 well plate. Agar arrays were printed using 2 uL and 4 uL drops. A time series of images was collected for 72 hrs. The data used to generate this figure are in Additional file 2. (a) From the final time point, spot intensity is plotted against spot area (scale to left), and normalized spot area is also plotted (scale to right). (b) Averaged data from all 96 cultures (4 μL drop array) are plotted for normalized and non-normalized spot intensities. Standard deviation bars show the effect of spot area normalization on measurement variation across time. (c) To further see the effect of normalization, arrays made with 2 μL and 4 μL spots from the same starting culture were analyzed. The averaged data from each set of 96 cultures, normalized and non-normalized, were plotted vs. time. (d) To observe the effect of spot area normalization on the MSR and AUGC, non-normalized and normalized values for each CPP were plotted vs. spot area.