Stankiewicz M, Shaw R J, Jonas W E, Cabaj W, Grimmett D J, Douch P G
Wallaceville Animal Research Centre, Upper Hutt, New Zealand.
Int J Parasitol. 1994 Apr;24(2):307-9. doi: 10.1016/0020-7519(94)90045-0.
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.
1至2岁的罗姆尼羊，通过至少三次感染80,000 - 100,000条蛇形毛圆线虫幼虫的驱虫简化感染程序进行免疫后，通常会产生大量肠道黏膜肥大细胞/球状白细胞（MMC/GLs）。在分离这些细胞时，强调了将肠道保持在37摄氏度、用二硫苏糖醇去除黏液、酶消化分散以及谨慎的体外处理程序以最大化细胞活力的重要性。通过使用Percoll不连续梯度从大多数污染细胞中分离出MMC/GLs。在60/100%Percoll界面收集的MMC/GLs通过补体包被的尼龙棉柱以去除污染的嗜酸性粒细胞。有活力的MMC/GLs能够在伴刀豆球蛋白A存在的情况下在体外生长，并在培养中存活长达30天。用金胺O染色后，通过紫外光显微镜很容易识别MMC/GLs。
