Supplementary Figure 14: Influence of inappropriate reference patterns.

(a,b) Reference patterns, in magenta / green / blue as used in Fig. 4 (3x ATTO 488), and in dark green and cyan inappropriate patterns generated for phalloidin/F-actin and EdU/DNA, respectively, from an older sample. The two patterns do not comply with all the requirements described in the Online Methods, i.e. the samples were prepared some days earlier than the multi-stained sample, possibly resulting in different staining conditions and sample decomposition. In addition they were measured one day after the multi-stained sample, possibly under slightly different optical and electronical settings. (c–f) Composite image (Golgi, F-actin, DNA) using an improper pattern for phalloidin/F-actin together with suited patterns for giantin/Golgi and EdU/DNA as used in Fig. 4. (g–i) Difference image to Fig. 4 (Golgi, F-actin, DNA), where green indicates an increase and magenta a decrease in signal. Numbers in brackets represent the photon counts of the brightest pixels. The mean of the absolute value of the difference is 125.6, 131.1, and 46.3 photons per pixel (g–i). (j–m) Composite image (Golgi, F-actin, DNA) using an improper pattern for EdU/DNA together with suited patterns for giantin/Golgi and phalloidin/F-actin as used in Fig. 4. (n–p) Difference image to Fig. 4 (Golgi, F-actin, DNA). The mean of the absolute value of the difference is 18.7, 134.5, and 120.5 photons per pixel (n–p). (q) Mere intensity image. Scale bars, 10 µm.