Automated method for isolation of adrenal medullary chromaffin cells from neonatal porcine glands.
An automated method for the isolation of neonatal porcine adrenal chromaffin cells is described. Adrenal chromaffin cells are potentially useful for therapeutic transplantation, but current isolation methodology suffers from labor intensiveness and variability in yield and viability due to imprecision of manual techniques, enzyme purity, and gland age and species. The described approach utilizes an adaptation of an automated procedure previously described for isolation of pancreatic islets. Results from neonatal porcine adrenal glands revealed consistent cell yields with high (approximately 99%) viability. Catecholamine assays showed that the resultant cultures continue to produce and secrete norepinephrine and epinephrine. Immunocytochemical analysis indicated that the majority of cells in the preparation are chromaffin cells and adrenal cortical cells. The procedure meets the following requirements: 1) minimal traumatic action on the adrenal chromaffin cells, 2) continuous digestion in which the adrenal cells that are progressively liberated can be saved from further mechanical action, 3) minimal human intervention in the digestion process, and 4) high yield and viability of the isolated adrenal chromaffin cells.
Vizzardelli
C
C
Diabetes Research Institute and The Miami Project to Cure Paralysis, University of Miami School of Medicine, FL 33136, USA.
Potter
E D
ED
Berney
T
T
Pileggi
A
A
Inverardi
L
L
Ricordi
C
C
Sagen
J
J
D000313:Adrenal Medulla / Q000166:cytology*; D000818:Animals; D000831:Animals, Newborn; D001331:Automation; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000166:cytology* / Q000378:metabolism; D004837:Epinephrine / Q000096:biosynthesis / Q000378:metabolism; D009638:Norepinephrine / Q000096:biosynthesis / Q000378:metabolism; D013552:Swine
D009638:Norepinephrine; D004837:Epinephrine