The use of a new dextran gradient medium for rapid isolation of functionally intact neonatal rat pancreatic islets.
The use of a new density gradient medium, Dextran M 70, is described for isolation of islets of Langerhans from collagenase digested pancreases of neonatal Wistar rats. Centrifugation in continuous as well as in discontinuous gradients of a less expensive dextran with a mol. wt. of 70,000 was performed and the results were compared with those obtained with Ficoll gradients. About 100 islets free from exocrine acinar parenchyma were obtained from each neonatal rat pancreas. There were no differences in glucose-stimulated insulin release and in potentiation of glucose-stimulated insulin release by the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX) between the islets isolated by centrifugation in Dextran M 70 or Ficoll 400 and those directly harvested from the pancreatic digest without density media. These data demonstrate that high numbers of pancreatic islets can be rapidly isolated in dextran gradients without any impairment of their functional integrity. Dextran M 70 gradients can be readily formed as continuous or 4-step discontinuous gradients without special osmotic compensators and its maximum density of 1.097 g/ml at a concentration of 23% (w/w) seems to be sufficient for the purification of other cell types too.
B
B
Hehmke
K D
KD
Kohnert
R
R
Odselius
D000818:Animals; D000831:Animals, Newborn; D002499:Centrifugation, Density Gradient / Q000379:methods*; D003911:Dextrans; D005362:Ficoll; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000502:physiology; D003012:Microbial Collagenase; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains
D003911:Dextrans; D007328:Insulin; D005362:Ficoll; D003012:Microbial Collagenase