Abstract:
:A method has been developed for the purification of single A, B and D cells and for the isolation of coupled islet cells. Isolated rat islets were dissociated by repeated pipetting in the presence of trypsin and ethylene glycol bis (beta-aminoethyl ether) N,N' tetra-acetic acid (EGTA), and then filtered through nylon and Percoll; the cell preparation consisted of 70% single and 30% coupled cells. Sizing of the cells led to the recognition of a small-islet cell population (35%; cell volume 200-600 micrometer 3) composed of A and D cells, and a large-islet cell population (65%; cell volume 600-1500 micrometer 3) identified as B cells. Differences in sedimentation velocity formed the basis for the islet cell separation by counterflow elutriation. Single islet cells eluted prior to coupled cells and were distributed over A and D cell-enriched fractions I and II (65% A, 25% B, 10% D) and the B cell-enriched fraction III (93% B). The slightly different densities of A (d = 1.068), B (d = 1.065) and D (d = 1.070) cells allowed a subsequent purification by density gradient centrifugation resulting in a final 10- to 30-fold enrichment in either A, B or D cells. Most coupled islet cells were recovered in fraction IV, occurred mainly as doublets and were composed of 90% B cells and 7% D cells; the multiple pseudopods, which characterize isolated D cells, might contribute to the coupling tendency of the D cells. It is concluded that the purified A, B and D cell fractions and the coupled islet cell preparations offer a direct approach to the study of individual islet cell types and their intercellular communication.
journal_name
Diabetologiajournal_title
Diabetologiaauthors
Pipeleers DG,Pipeleers-Marichal MAdoi
10.1007/BF00257436subject
Has Abstractpub_date
1981-06-01 00:00:00pages
654-63issue
6eissn
0012-186Xissn
1432-0428journal_volume
20pub_type
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