Storage of porcine pancreatic digest prior to islet purification. The benefits of UW solution and the roles of its individual components.
The aim of this study was to establish the beneficial effect of storage of pancreatic digest in University of Wisconsin solution on porcine islet purification, the mechanism of this effect, and the components of UW responsible. Ten porcine pancreata were collagenase-digested, and samples of digest were washed and stored for 1 hr in either UW or minimum essential medium at 4 degrees C, prior to separation on continuous linear density gradients of bovine serum albumin. Samples of digest from a further ten pancreata were similarly treated, comparing storage in MEM, UW, and five solutions varying in lactobionate:chloride ratio and raffinose content. The purity of the islet preparations and the densities of islets and exocrine tissue were determined from insulin and amylase assay of aliquots aspirated from these gradients. Washing and storage of digest in UW markedly improved islet purity, compared with MEM, due to an increase in the density of exocrine tissue. Exocrine tissue density following storage was dependent upon the control of acinar cell volume, rather than exocrine enzyme discharge, and was determined primarily by the chloride:lactobionate ratio of the storage solution. Raffinose was of little additional benefit, while the beneficial effect of UW was greater than that due to its lactobionate and raffinose content alone. In conclusion, inadequate purification of islets results from exocrine tissue swelling. This swelling is reduced by storage of the pancreatic digest in UW solution, due primarily to the replacement of chloride by lactobionate in UW.
Chadwick
D R
DR
Department of Surgery, Leicester Royal Infirmary, United Kingdom.
Robertson
G S
GS
Rose
S
S
Contractor
H
H
James
R F
RF
Bell
P R
PR
London
N J
NJ
D000241:Adenosine / Q000737:chemistry; D000493:Allopurinol / Q000737:chemistry; D000818:Animals; D002499:Centrifugation, Density Gradient; D005978:Glutathione / Q000737:chemistry; D007328:Insulin / Q000737:chemistry; D007515:Islets of Langerhans / Q000166:cytology*; D019188:Organ Preservation Solutions; D010179:Pancreas / Q000737:chemistry / Q000166:cytology; D011887:Raffinose / Q000737:chemistry; D013552:Swine; D014021:Tissue Preservation / Q000379:methods*
D007328:Insulin; D019188:Organ Preservation Solutions; C055206:University of Wisconsin-lactobionate solution; D000493:Allopurinol; D005978:Glutathione; D000241:Adenosine; D011887:Raffinose