(a) Transfer molten agarose to slide containing plastic spacers (shown in gray). (b) Quickly add second slide to form pad. (c) After agarose has cooled, remove upper slide and spacers. (d) Add polystyrene nanoparticles. (e) Add washed worm(s). (f) Add coverslip. The worms are now ready for imaging. (g) Seal coverslip with wax if necessary. (h) To recover animals, lift coverslip upwards without sliding.

(a) Fluorescence of AFD soma in a nanoparticle-immobilized Pgcy-8::YC3.60 worm. Centroid of cell body indicated by crosshairs. Scale bar: 20 µm. (b) NP: Mean absolute displacement of AFD neuron cell body during 10 s intervals in animals immobilized with 0.1 µm diameter nanoparticles, as a function of agarose concentration in pad. NGM: no beads. (c) Mean absolute displacement of buccal cavity during 10 s intervals for adult worms on 10% agarose pads and varied bead diameters; L1, L3, and adult (Ad) worms immobilized with 0.1 µm beads on 10% agarose pads.

(a) Adult hermaphrodite in NGM buffer. (b) Same animal during immobilization on 10% agarose pad. Arrows indicate measurement of dorso-ventral width. Scale bars: 100 µm. (c) Dorso-ventral width as a function of agarose concentration for 0.1 µm diameter nanoparticle-immobilized worms (n>5 for each point).

Locomotory frequency of freely swimming worms and worms in contact with 0.5% agarose pads, in the presence (NP) and absence (NGM) of nanoparticles. n = 30 for each group. **p<0.001.

a) Images from a continuously immobilized C. elegans, showing an ALM neuron severed 20 µm from the cell soma (top panel) immediately following laser axotomy and (bottom panel) after 10 hr. Arrow indicates lesion point. b) Average regenerative outgrowth of ALM neurons severed at the indicated distances. Outgrowth was measured from images taken at 10 hr post surgery and categorized as initiating from the lesion point or from the cell soma (* indicates p<0.05 by Student’s t-test). c) Average FRET signals measuring cellular calcium response to laser axotomy. At each time point, two channel fluorescent FRET images of the target neuron where averaged across the cell soma and along the axon within 5 µm of the lesion point to measure intracellular calcium levels in those regions. Arrows indicate time of laser damage (t = 0 s). Shaded regions indicate standard error on the mean at each time point. n indicates number of worms assayed.