title: Microbial surveillance of human islet isolation, in vitro culture, and cryopreservation.
abstract: The aim of these studies was to investigate microbiological contamination during the isolation and preservation of islets of Langerhans in a low-temperature tissue bank. Islets were isolated from the pancreases of 117 organ donors, then cryopreserved. In the initial 47, microbial culture was completed only after thawing: Enterobacter cloacae (n = 4) and gram-negative bacilli (n = 9) were isolated for a positive culture rate of 27.6%. It was not possible to ascertain the source of the contaminants, since cultures were taken only at the conclusion. A total of 70 consecutive pancreases were then subjected to islet isolation and cryopreservation while prospectively culturing at these steps: step 1, from the pancreas transport media; step 2, after intraductal perfusion of collagenase; step 3, after dissociation; step 4, after purification; step 5, following a 6-48-h in vitro culture; step 6, post-thaw; step 7, after dilution of the cryoprotective agent; and step 8, after final 48-h in vitro culture. The transport fluid was infected with aerobic gram-negative (n = 6), gram-positive (n = 5), and yeast microorganisms (n = 2) for an overall step 1 contamination rate of 19%. These contaminants were found in 9% of our local program vs. 26% from distantly procured pancreases. Contaminants at subsequent steps were 10% (step 2), 9% (step 3), so that only 3% were infected at the conclusion of isolation (step 4), and 0% after initial culture (step 5). Amongst pancreases that were initially sterile, new contaminants could be identified in 12% at step 2 and 8% at step 3; however, these also became undetectable.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Surgery, University of Alberta, Edmonton.
authors.forename: J R
authors.initials: JR
authors.lastname: Lakey
authors.forename: R V
authors.initials: RV
authors.lastname: Rajotte
authors.forename: G L
authors.initials: GL
authors.lastname: Warnock
mesh_terms: D001419:Bacteria / Q000302:isolation & purification*; D015925:Cryopreservation; D046508:Culture Techniques; D006801:Humans; D007515:Islets of Langerhans / Q000382:microbiology*; D016381:Islets of Langerhans Transplantation; D011446:Prospective Studies; D012189:Retrospective Studies; D014015:Tissue Banks