Primary cultures of embryonic cells from the snail Biomphalaria glabrata

View More View Less
  • 1 Stanford University School of Medicine, Department of Community and Prenentive Medicine, Stanford, California 94305

Whole egg masses or separated capsules containing trochophore-stage embryos of Biomphalaria glabrata were axenized in dilute hypochlorite with or without antibiotics. Embryos were extracted in snail saline and teased into fragments which were dissociated (generally with trypsin) and seeded into media. Of numerous formulations used, 4 were equally good: 3 are based upon diluted medium 199 or NCTC-135, with additions, and the other was formulated from analyses of hemolymph and published values; 3 are used under 5% CO2 and the other in air. Osmolalities vary from 116 to 220 mOsm/Kg H20. Fetal bovine serum is added to all and appears necessary to sustain cell growth. Cultures were not enhanced by use of collagen coatings, plasma clots, or adult snail extracts. Snail hemolymph is toxic when fresh and ineffective after heat inactivation. Five cell types were seen: fibroblast-like, principally in the first 2 weeks; epithelial-like, sometimes making sheets or monolayers; attenuated contractile muscle cells which sometimes became broad and binucleate; macrophage-like; and hemocytes. Numerous mitotic figures have been observed in aceto-orcein squash preparations, especially after incubation with colcemid. Cultures were usually healthy until terminated up to 6 weeks old.