The evaluation of neutral density separation utilizing Ficoll-sodium diatrizoate and Nycodenz and centrifugal elutriation in the purification of bovine and canine islet preparations.
This study evaluates the potential of a neutral density separation technique using iodinated gradient media in combination with Ficoll or alone at two different densities for the purification of bovine and canine islets isolated by a continuous digestion-filtration technique. In addition, centrifugal elutriation is applied to the purification of isolated bovine islets. A single density layer of Ficoll-sodium diatrizoate or Nycodenz at densities of 1.080 g/ml or 1.085 g/ml resulted in an recovery of 47.4% to 77.4% bovine islet volume with bovine islet purities ranging from 43.5% to 49.0%. The use of a single density layer of Ficoll-sodium diatrizoate at 1.080 g/ml and Nycodenz at 1.085 g/ml resulted in recoveries of 86.4% and 56.2% canine islet volume with canine islet purities of 58% and 63.5%, respectively. The bovine islet enriched fraction (flow rates 50-90 ml/min at a rotor speed of 200 rpm) obtained by centrifugal elutriation was 20.0 +/- 3.5% pure, the average recovery of bovine islet volume obtained by centrifugal elutriation was 59.0 +/- 10.7%. It is suggest that a further refinement of factors like osmolality, viscosity, pH, ionic composition and temperature of iodinated density gradient media may allow continued improvement of islet purity and recovery obtained by the simple neutral density separation technique.
Hering
B J
BJ
Medizinische Klinik III, Justus Liebig Universität, Giessen, Germany.
Muench
K P
KP
Schelz
J
J
Amelang
D
D
Heitfeld
M
M
Bretzel
R G
RG
Bonath
K
K
Federlin
K
K
D000818:Animals; D002417:Cattle; D002469:Cell Separation / Q000379:methods*; D002498:Centrifugation / Q000379:methods*; D002499:Centrifugation, Density Gradient / Q000379:methods*; D004247:DNA / Q000032:analysis; D003973:Diatrizoate; D004285:Dogs; D005260:Female; D005362:Ficoll; D007472:Iohexol; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D008991:Monitoring, Physiologic; D014183:Transplantation, Heterologous
D003973:Diatrizoate; D005362:Ficoll; D007472:Iohexol; D004247:DNA