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Sorting of pancreatic islet cell subpopulations by light scattering using a fluorescence-activated cell sorter

Nielsen, D A ; Lernmark, A LU orcid ; Berelowitz, M ; Bloom, G D and Steiner, D F (1982) In Diabetes 31(4 Pt 1). p.299-306
Abstract

Methods have been developed for the preparation of suspensions of viable rat pancreatic islet cells and their analysis and sorting in the fluorescence-activated cell sorter (FACS III or IV). Histograms of cell number versus light scattering in a near forward angle (1-15 degrees) demonstrated that viable islet cells produce a broad peak that is distinctly separated from the peaks generated by exocrine cells, erythrocytes, and nonviable cells. Electron microscopic examination and radioimmunoassay of hormone content in fractions collected across the peak showed that glucagon-containing (A) cells scatter less intensely and are concentrated within the left side of the islet cell peak, while somatostatin-containing (D) cells are localized to... (More)

Methods have been developed for the preparation of suspensions of viable rat pancreatic islet cells and their analysis and sorting in the fluorescence-activated cell sorter (FACS III or IV). Histograms of cell number versus light scattering in a near forward angle (1-15 degrees) demonstrated that viable islet cells produce a broad peak that is distinctly separated from the peaks generated by exocrine cells, erythrocytes, and nonviable cells. Electron microscopic examination and radioimmunoassay of hormone content in fractions collected across the peak showed that glucagon-containing (A) cells scatter less intensely and are concentrated within the left side of the islet cell peak, while somatostatin-containing (D) cells are localized to the far right side, indicating a higher intrinsic light scattering property of the D-cells. The more abundant insulin-containing (B) cells define the center of the islet cell peak. Sodium dodecyl sulfate slab gel electrophoresis and radioautography of 35S-methionine labeled cellular proteins confirmed that sorted cells are viable. Cells from the far left region contained increased amounts of labeled 18 Kd proglucagon and its 13-Kd and 10-Kd conversion intermediates, while cells from the right side were relatively enriched in labeled 12.4 Kd prosomatostatin. These results demonstrate that intrinsic light scattering alone can be used to prepare A- or D-cell enriched fractions from islets for biochemical analysis.

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author
; ; ; and
publishing date
type
Contribution to journal
publication status
published
keywords
Animals, Cell Separation/methods, Chickens, Erythrocytes, Exocrine Glands/cytology, Flow Cytometry/methods, Glucagon/analysis, Insulin/analysis, Islets of Langerhans/cytology, Male, Microscopy, Electron, Rats, Rats, Inbred Strains, Somatostatin/analysis
in
Diabetes
volume
31
issue
4 Pt 1
pages
299 - 306
publisher
American Diabetes Association Inc.
external identifiers
  • scopus:0020054058
  • pmid:6130019
ISSN
0012-1797
DOI
10.2337/diab.31.4.299
language
English
LU publication?
no
id
0f8adf1d-e3c7-4b52-8c2e-5711f3a3c184
date added to LUP
2021-09-14 16:35:22
date last changed
2024-03-13 08:06:15
@article{0f8adf1d-e3c7-4b52-8c2e-5711f3a3c184,
  abstract     = {{<p>Methods have been developed for the preparation of suspensions of viable rat pancreatic islet cells and their analysis and sorting in the fluorescence-activated cell sorter (FACS III or IV). Histograms of cell number versus light scattering in a near forward angle (1-15 degrees) demonstrated that viable islet cells produce a broad peak that is distinctly separated from the peaks generated by exocrine cells, erythrocytes, and nonviable cells. Electron microscopic examination and radioimmunoassay of hormone content in fractions collected across the peak showed that glucagon-containing (A) cells scatter less intensely and are concentrated within the left side of the islet cell peak, while somatostatin-containing (D) cells are localized to the far right side, indicating a higher intrinsic light scattering property of the D-cells. The more abundant insulin-containing (B) cells define the center of the islet cell peak. Sodium dodecyl sulfate slab gel electrophoresis and radioautography of 35S-methionine labeled cellular proteins confirmed that sorted cells are viable. Cells from the far left region contained increased amounts of labeled 18 Kd proglucagon and its 13-Kd and 10-Kd conversion intermediates, while cells from the right side were relatively enriched in labeled 12.4 Kd prosomatostatin. These results demonstrate that intrinsic light scattering alone can be used to prepare A- or D-cell enriched fractions from islets for biochemical analysis.</p>}},
  author       = {{Nielsen, D A and Lernmark, A and Berelowitz, M and Bloom, G D and Steiner, D F}},
  issn         = {{0012-1797}},
  keywords     = {{Animals; Cell Separation/methods; Chickens; Erythrocytes; Exocrine Glands/cytology; Flow Cytometry/methods; Glucagon/analysis; Insulin/analysis; Islets of Langerhans/cytology; Male; Microscopy, Electron; Rats; Rats, Inbred Strains; Somatostatin/analysis}},
  language     = {{eng}},
  number       = {{4 Pt 1}},
  pages        = {{299--306}},
  publisher    = {{American Diabetes Association Inc.}},
  series       = {{Diabetes}},
  title        = {{Sorting of pancreatic islet cell subpopulations by light scattering using a fluorescence-activated cell sorter}},
  url          = {{http://dx.doi.org/10.2337/diab.31.4.299}},
  doi          = {{10.2337/diab.31.4.299}},
  volume       = {{31}},
  year         = {{1982}},
}