A technique for the isolation and purification of viable mucosal mast cells/globule leukocytes from the small intestine of parasitised sheep.
Romney sheep, 1-2 years old, immunized by at least three anthelmintic abbreviated infections of 80-100,000 Trichostrongylus colubriformis larvae usually produced high numbers of intestinal mucosal mast cells/globule leukocytes (MMC/GLs). In isolating these cells, the importance of maintaining the intestine at 37 degrees C, removal of mucus with dithiothreitol, enzymatic dispersion and careful in vitro handling procedures for maximising cell viability are emphasised. The MMC/GLs were separated from most contaminant cells by using a Percoll discontinuous gradient. MMC/GLs collected at the 60/100% Percoll interface were passed through a complement coated nylon wood column to remove the contaminating eosinophils. Viable MMC/GLs were able to grow in vitro in the presence of Concanavalin A and survive in culture for up to 30 days. The MMC/GLs were readily identified by ultraviolet light microscopy after staining with auramine O.
Stankiewicz
M
M
Wallaceville Animal Research Centre, Upper Hutt, New Zealand.
Shaw
R J
RJ
Jonas
W E
WE
Cabaj
W
W
Grimmett
D J
DJ
Douch
P G
PG
D000818:Animals; D002469:Cell Separation; D002499:Centrifugation, Density Gradient; D007413:Intestinal Mucosa / Q000473:pathology; D007421:Intestine, Small / Q000473:pathology*; D007962:Leukocytes; D008407:Mast Cells; D012756:Sheep; D012757:Sheep Diseases / Q000276:immunology*; D014253:Trichostrongylosis / Q000276:immunology / Q000662:veterinary*