Fig. 6

Fluorescence comparison of mTagBFP2, Electra1 and Electra2 in small model organisms. (a) Whole body images of representative C. elegans worms co-expressing mTagBFP2, Electra1 and Electra2 with mNeonGreen under pan-neuronal promoter tag168 and mScarlet in most somatic cells. Higher magnification images of neuronal ring under blue (shown in grey), green (mNeonGreen), red channel (mScarlet) and merged. Imaging conditions: 405 nm excitation for BFPs, 430–470 nm emission; 488 nm excitation for mNeonGreen, 500–540 nm emission; 561 nm excitation for mScarlet, 570–620 nm emission. The dynamic range of all images was adjusted independently to facilitate visualization. (b) Blue-to-green fluorescence ratio comparison in C. elegans neuronal ring (n = 9 worms for each protein; Kruskal Wallis ANOVA p-value = 1.29e-4; post-hoc two sample Kolmogorov-Smirnoff p-values shown in graph; Supplementary Table S7). (c) Representative whole overview image of 4dpf zebrafish expressing each BFP throughout brain and spinal cord (left). Higher magnified images of blue, red, and merged fluorescence in hindbrain (middle) and spinal cord (right) neurons. Transient co-expression of each BFP and mScarlet was induced by injection into zebrafish embryos with a construct carrying each BFP-P2A-mScarlet whose expression is under the control of pan-neuronal nbt promoter. Imaging conditions: Blue channel: excitation 405 nm laser, emission 420–480 nm; red channel: excitation 561 nm laser, emission 565–606 nm). The dynamic range of all images was adjusted independently to facilitate visualization. (d) Blue-to-red fluorescence ratio comparison in zebrafish hindbrain expressing mTagBFP2, Electra1, Electra2 (n = 116, 117, 116 neurons from 4 independent zebrafish; Kruskal–Wallis ANOVA p-value = 1.98e-19; post-hoc two sample Kolmogorov-Smirnoff p-values shown in graph; Supplementary Table S8; imaging conditions for BFPs: same as in (a)). (e) Blue-to-red fluorescence ratio comparison in zebrafish spinal cord neurons expressing mTagBFP2, Electra1, Electra2 (n = 113, 118, 117 neurons from 4 independent zebrafish; Kruskal–Wallis ANOVA p-value = 1,2e-8; post-hoc two sample Kolmogorov-Smirnoff p-values shown in graph; Supplementary Table S8) with mScarlet. Imaging conditions for BFPs: same as in (c). (f)Time-dependent blue fluorescence measurement for mTagBFP2, Electra1, Electra2 (n = 40, 40, 40, respectively from 4 independent zebrafish) in spinal cord neurons. Imaging conditions of BFPs: Blue channel: excitation 405 nm laser, emission 420–464 nm.