Three-dimensional imaging of live Caenorhabditis elegans commonly relies on sequential z-stack acquisition, which can be slow and susceptible to movement and photobleaching. Here, we used light-field microscopy to capture dye and delivery-dependent fluorescence patterns across intact adult worms in a single exposure per channel. With a 40x/1.20 NA water-immersion objective, each reconstructed dataset contained 121 axial slices spanning 109 {micro}m at 0.91 {micro}m spacing. In a representative adult, nematode signal extended from 40.88 to 85.4 {micro}m, corresponding to approximately 44.5 {micro}m of captured axial depth without sequential z scanning. Three fluorescence channels were acquired with a summed exposure time of 112 ms across a 685 x 1010 {micro}m field of view. FM1-43 labelled intestinal, cuticular and vesicular structures, FM4-64 highlighted ingested bacteria and intestinal compartments, and Nile Red revealed vesicular and broader whole-organism fluorescence. Intrinsic blue and green autofluorescence signals were also resolved. These observations demonstrate rapid single-shot acquisition of whole-organism three-dimensional fluorescence information and show how delivery route influences the spatial distribution of commonly used fluorescent probes in C. elegans.
Markus, R., Rea, E., Berfi, N. A., Taresco, V., Hudson, C., White, D. R. A., Chauhan, V. M.
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