Table 2 Specific challenges when imaging pLLP morphogenesis.

pLL morphogenesis lasts about 30 h: Embryos need to be immobilized but still survive and grow over a longtime period

Use of low concentration of LMPA containing Tricaine

pLLP migrates over almost 2 mm so the area to image is 2 to 3 times longer than one field of view (FOV):

(i) If the embryos are not mounted parallel to the coverslip, the size of Z-stack increases significantly.

(ii) The contra-lateral pLLP eventually enters the Z-volume and introduce interfering signal in post-processing Z-projections.

Multiple FOV need to be imaged and stitched

The use of μ-wells with a defined shape orients the tails parallel to the coverslip (see Fig. 2a)' to both reduce the size of Z-stacks and prevent the interference of the contra-lateral pLLP.