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Nonenzymic in vitro isolation of perinatal islets of Langerhans

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Summary

We have developed a method to circumvent the use of exogenous proteolytic enzymes in the isolation of islets of Langerhans from the perinatal rodent pancreas. Advantage is taken of the propensity of fibroblastlike cells to attach and migrate on polystyrene at low-serum concentrations (5%). In contrast, at this serum level, rat islet epithelial cells tend not to adhere to the substrate. At 3 d of culture, islets are visible at the edges of the explants. With further fibroblast outgrowth the majority of islets are freefloating by 7 d. Simple agitation of the medium and centrifugation yields approximately 50 μg of islet tissue per perinatal pancreas. Further purification of the islets can be obtained by subculture. Rat islets can be maintained in this manner for several months in Medium F12 supplemented with 25% horse serum in an atmosphere of 5% CO2 and air at 37° C. Hormone content of the islet tissue remains constant during prolonged subculture and such islets continue to exhibit appropriate insulin and glucagon responses to glucose and theophylline. The morphological integrity of the endocrine cells within the cultured islets was confirmed by immunocytochemistry and ultrastructural study. Nonendocrine cells are not identifiable within the long-term cultured islets.

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Author notes

  1. Bradley A. Schulte

    Present address: Department of Pathology, Medical University of South Carolina, 171 Ashley Avenue, 29403, Charleston, South Carolina

  2. Janet R. Serie

    Present address: The College of St. Catherine, 2004 Randolph Avenue, 55105, St. Paul, Minnesota

Authors and Affiliations

  1. Department of Anatomy, University of Minnesota, 55455, Minneapolis, Minnesota

    Orion D. Hegre, Sue Marshall, Bradley A. Schulte, Gregg E. Hickey, Frank Williams, Robert L. Sorenson & Janet R. Serie

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  1. Orion D. Hegre
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  2. Sue Marshall
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  3. Bradley A. Schulte
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  4. Gregg E. Hickey
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  5. Frank Williams
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  6. Robert L. Sorenson
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  7. Janet R. Serie
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Additional information

This research was supported in part by Grants AM 19899 and HD 00412 from the National Institutes of Health, Bethesda, MD, and grants from the American Diabetes Association, Minnesota Affiliate.

Portions of this work were presented at the Thirty-third Annual Meeting of the Tissue Culture Association, held in San Diego, California, June 6–10, 1982.

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Hegre, O.D., Marshall, S., Schulte, B.A. et al. Nonenzymic in vitro isolation of perinatal islets of Langerhans. In Vitro 19, 611–620 (1983). https://doi.org/10.1007/BF02619574

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