Immunocytochemistry
The epithelial origin of the isolated BOECs was
confirmed by subjecting the cells to immunocytochemical
staining for the expression of cytokeratins using monoclonal
mouse antihuman cytokeratin antibodies (clone AE1/
AE3), biotinylated goat antimouse antibodies, streptavidin–
horseradish peroxidase, and AEC substrate-chromogen
(Dako, Denmark) according to the manufacturer’s instructions.
Bovine oviductal epithelial cells were grown in
four-well culture plates: (1) as freshly retrieved BOECs, (2)
as first-passage BOECs after a 7-day culture period in a
culture flask (to increase cell purity), trypsinized, and
reseeded on glass inlets, and (3) as freshly retrieved BOECs
cultured as monolayers on hanging inserts in a PCC system.
Cytokeratin-stained cells were compared to nonstained
cells, and a fibroblast culture of fetal bovine cartilage was
used as a negative control, and nonspecific binding