The cell blot assay in analysis of rat anterior pituitary cell secretion.
We have investigated the efficacy of the cell blot assay in analysis of the secretion of hormones and peptides from rat anterior pituitary cells. The dissociated cells are cultured on pieces of translucent polyvinylidene difluoride membrane, on which their secretory products are adsorbed and subsequently immunostained. The area and integrated optical density of the stained 'halo' surrounding individual cells is measured by microscopical image processing and the values for basal secretion of a particular hormone or peptide are compared with those after application of secretagogues or inhibitors. Our experiments tested established responses of dissociated rat anterior pituitary cells; in general, the results were as expected. Double immunoenzymatic staining could be used to show secretion of two products from the same or different cells in one preparation, and immunofluorescence with fluorescein- and/or rhodamine-labelled antibodies could be used instead of enzyme-linked immunolabelling. Optimal dilutions of immunoreagents were much higher than those used for immunocytochemistry on tissue sections. Although the cell blot assay does not provide absolute quantification, since some of the secreted product escapes into the medium, it is a relatively easy and economical way for morphologists to compare secretion from individual cells under varying conditions.
Cimini
V
V
Department of Human Anatomy, Medical School, Federico II University of Naples, Italy.
Van Noorden
S
S
Mahadeva
H
H
Polak
J M
JM
D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D003078:Colchicine / Q000494:pharmacology; D003345:Corticosterone / Q000494:pharmacology; D003346:Corticotropin-Releasing Hormone / Q000494:pharmacology; D004298:Dopamine / Q000494:pharmacology; D019004:Galanin; D007150:Immunohistochemistry; D008297:Male; D009479:Neuropeptides / Q000032:analysis* / Q000378:metabolism; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology; D010903:Pituitary Gland, Anterior / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D011388:Prolactin / Q000378:metabolism; D051381:Rats; D012636:Secretory Rate / Q000187:drug effects; D013268:Stimulation, Chemical; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism; D014667:Vasopressins / Q000494:pharmacology
D009479:Neuropeptides; D010455:Peptides; D014667:Vasopressins; D014660:Vasoactive Intestinal Peptide; D013973:Thyrotropin-Releasing Hormone; D019004:Galanin; D000324:Adrenocorticotropic Hormone; D011388:Prolactin; D003346:Corticotropin-Releasing Hormone; D003078:Colchicine; D004298:Dopamine; D003345:Corticosterone