Established methods for isolating intraepithelial lymphocytes (IEL) from mouse small intestine were modified to facilitate the isolation of IEL from rainbow trout gut epithelium. A 1h incubation in 1m
MDTT/EDTA was required to maximise the separation of the epithelial compartment from the underlying lamina propria. Epithelial cell content was reduced by filtration on nylon wool columns followed by centrifugation on a 40 % Percoll cushion. Most IEL pelleted through the 40 % Percoll [high density (HD) IEL] although significant numbers banded out at the medium/40 % Percoll interface [low density (LD) IEL]. The majority (
90 % ) of HD IEL were<12·5μm in size, had a pleiomorphic nucleus and obvious mitochondria. LD IEL were larger cells, containing more cytoplasm with abundant ribosomes. Neither HD nor LD IEL appeared to contain cytotoxic granules. Despite this, in a non-radioactive cytotoxicity assay (LDH release) with EL4 mouse thymoma cells as targets, significant killing was observed by both the HD and LD IEL.