Fig. 3. Histology and cytochemistry of extirpated buds and degenerating zooids isolated from Botryllus schlosseri colonies. (A–C) An epithelial monolayer that developed from an isolated zooid in mid-takeover, 14-day culture period (A, x200; B, x400). At 3 weeks incubation (C, x200), the monolayer has started to deteriorate. TUNEL assay did not reveal positive nuclei. (D) An epithelial monolayer originated from stage `C' bud (x200) at the deteriorating stage, 4 weeks in culture. TUNEL assay performed on this monolayer did not reveal positive nuclei. (E,F) Apoptotic cells in histological section carried out on the extirpated stage `D' and TUNEL-stained zooid (E, x200); also stained for general morphology by Azan (F, x200). (G,H) TUNEL-positive cells (arrowhead) in histological sections of 5-week old cultured buds, counterstained by Methyl Green (G, x1000; H, whole bud, x200). cb, cell boundary; dv, disintegrated visceral organs; e, epithelium; me, monolayer edge; s, stomach; v, vacuole; vo, visceral organ; n, nucleus. Scale bars, 50 µm.