JCS02625 Supplementary Material

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• Supplemental Figure 1 -

  Fig. S1. Measuring calcium signals in single cells of the zebrafish embryo (A) Optical section, showing side view through the trunk of a dye-labelled embryo at 19 hours, with dorsal towards the top and anterior to the right. Muscle fibres, loaded with Oregon Green 488 BAPTA dextran and rhodamine dextran, in a position consistent with the nascent horizontal myoseptum (white box) were identified. Images were captured using confocal laser scanning microscopy (Zeiss 510) and Zeiss Fluar X20 objective (N.A. 0.75). (B) Changes in fluorescence were calculated as the average pixel intensity within user-defined regions drawn around the muscle fibre and plotted against time. A total of 200 images for each indicator were captured. All images were processed and analyzed using Zeiss software. Fluorescence-emission-intensity from Oregon Green 488 BAPTA dextran (green) and rhodamine dextran (red) report Ca²⁺-concentration-dependent and -independent changes, respectively. (C) Changes in the ratio of Oregon Green 488 BAPTA dextran versus rhodamine dextran were calculated from emission-intensity measurements. A Ca²⁺ transient was defined as an increase, at least twice that of the standard deviation, above the baseline that remained above this level for a minimum of two points but then returned to baseline. Images that contain the whole muscle fibre were taken every 380 ms over a two-minute period, and this was repeated every 15 minutes.