Suitability of different staining methods for the identification of isolated and cultured cells from guinea pig (Cavia aperea porcellus) stomach.
Cell suspensions from the guinea pig gastric mucosa were obtained using a pronase/collagenase isolation method, and cultured on Petri dishes in minimum essential medium at 37 degrees C. For proper identification of different gastric cell types in cytospots, cell suspensions or culture, selective staining methods were employed, modified and evaluated. Mucous cells and mucous neck cells were detected by use of lectins. Mucous cells were stained on cytospots and in primary cultures with lectins from peanut, Helix pomatia, Ulex europaeus, wheat germ, and from soybean. Vital chief cells in suspensions but not in culture, were selectively stained by Nile blue sulphate, brilliant cresyl blue or the fluorescence dye dihexyloxacarbocyanine iodide. Pepsinogen granules of isolated and cultured chief cells were detected with a polyclonal antibody against porcine pepsinogen. Isolated parietal cells were identified in cytospots by using acidophilic dyes (aurantia, eosin). In suspensions and in cultures vital parietal cells were identified by enzymatic detection of succinic dehydrogenase or carboanhydrase activity and by the vital stain Janus green. In cultures exclusively, parietal cells were additionally identified by the vital stain rhodamine. Cytochemically, they were identified with phalloidin by binding to actin filaments. Endocrine cells in the suspension were visualised immunocytochemically with antibodies directed against different amines or peptides. Fibroblasts and endothelial cells were identified after isolation and in primary culture with a vimentin antibody. Mast cells in suspension were either visualised by a histamine antibody or by metachromatic staining behaviour to toluidine blue, respectively. Endothelial cells in suspension or culture were distinguished from fibroblasts by endocytosis of acetylated low-density-lipoprotein. In conclusion, the developed methods are highly suitable to identify guinea pig gastric cells after isolation and follow up their fate in primary culture.
Giebel
J
J
Institute for General Pharmacology; Medical School Hanover, Germany.
Arends
H
H
Fanghänel
J
J
Cetin
Y
Y
Thiedemann
K U
KU
Schwenk
M
M
D000818:Animals; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D004396:Coloring Agents; D005753:Gastric Mucosa / Q000166:cytology*; D006168:Guinea Pigs; D007150:Immunohistochemistry; D010295:Parietal Cells, Gastric / Q000166:cytology
D004396:Coloring Agents