Supplementary Materials

Observing the cell in its native state: Imaging subcellular dynamics in multicellular organisms

Tsung-Li Liu, Srigokul Upadhyayula, Daniel E. Milkie, Ved Singh, Kai Wang, Ian A. Swinburne, Kishore R. Mosaliganti, Zach M. Collins, Tom W. Hiscock, Jamien Shea, Abraham Q. Kohrman, Taylor N. Medwig, Daphne Dambournet, Ryan Forster, Brian Cunniff, Yuan Ruan, Hanako Yashiro, Steffen Scholpp, Elliot M. Meyerowitz, Dirk Hockemeyer, David G. Drubin, Benjamin L. Martin, David Q. Matus, Minoru Koyama, Sean G. Megason, Tom Kirchhausen, Eric Betzig

Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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  • Supplementary Text
  • Figs. S1 to S20
  • Table S1
  • Captions for Movies S1 to S8
  • References

Images, Video, and Other Media

Movie S1
Volume rendering of 200 nm beads embedded in 1% agarose imaged with no correction (left) and with complete AO correction (detection, excitation and autofocus, right). The volume is rotated around the x axis showing its z and y projections (c.f., Fig 1).
Movie S2
Subcellular structure and dynamics in the spine of a zebrafish embryo 24 hpf expressing markers for the cell surface (mCardinal-membrane), trans-Golgi apparatus (mNeonGreen-GalT) and mitochondria (TagRFPt-cox8a). Part 1 of the movie compares volume renderings at a single time point using unprocessed data, deconvolved data without AO correction, and deconvolved data with AO correction. Part 2 compares, at a single time point, xy maximum intensity projections of unprocessed data, data with AO correction only, and data with AO and deconvolution Part 3 shows the dynamics of the organelles after AO correction plus deconvolution (c.f., Fig 1D).
Movie S3
Dynamics of CCPs and CCVs (green) relative to muscle cell membranes, including their t-tubules (red), in a 2μm slab through the tail of a zebrafish embryo 50-55 hpf. Both CCPs pinned to t-tubules and CCVs rapidly shuttling between t-tubules along the fiber axis are observed. (c.f., Fig. 2, Movie 3).
Movie S4
Movie S4. A large image volume assembled from 7 x 7 x 3 subvolumes in the tail of a zebrafish embryo 96 hpf comparing three stitched and deconvolved datasets: (top) no AO correction; (middle) AO correction from center tile applied to all tiles; and (bottom) independent correction applied to each tile (c.f., fig 4).
Movie S5
Volume rendered time series of zebrafish spine development from 30-34 hpf imaged at 30 min intervals. The excitation and detection path aberrations are shown on either side of the aberration corrected spine volume. The subset of tiles at a given time point where AO corrections are updated are highlighted with green boxes (c.f., Fig 4D).
Movie S6
Sagittal view of the migration of rostrocaudally projecting axons of newly differentiated neurons labeled by Autobow, imaged at 10.4 min intervals from 58 to 70 hpf (c.f., Fig. 6A-C).
Movie S7
Visualizing C. elegans AC invasion in vivo. Basement membrane (magenta) and ACspecific F-actin (green) in the C. elegans L3 stage somatic gonad and vulval epithelium, prior to the time of AC invasion, showing xy and xz orthoslices and volume rendered views before (left) and after (right) AO correction and deconvolution (c.f., fig S17).
Movie S8
Arabidopsis cotyledon epidermal cells expressing microtubule reporter p35S::GFPMBD, showing xy and xz orthoslices and volume rendered views before (left) and after (right) AO correction and deconvolution (c.f., fig. S18).