Preparation and characterization of plasma membrane-enriched fractions from rat pancreatic islets
(1976) In Journal of Cell Biology 71(2). p.606-623- Abstract
Methods have been developed for the isolation on a semi-micro scale of a plasma membrane-enriched fraction from rat islets of Langerhans. An important feature of these experiments is the use of 125I-labeled wheat germ agglutinin as a specific probe for plasma membrane-containing fractions. The partly purified plasma membrane fraction had a density in sucrose of about 1.10 and was enriched in the activities of 5′-nucleotidase, alkaline phosphatase, sodium-potassium, and magnesium- dependent ATPases and adenylate cyclase. It contained only very low levels of acid phosphatase, cytochrome c oxidase, insulin, and RNA. Further purification was hampered by the relatively small amounts of fresh plasma membrane material that could be obtained... (More)
Methods have been developed for the isolation on a semi-micro scale of a plasma membrane-enriched fraction from rat islets of Langerhans. An important feature of these experiments is the use of 125I-labeled wheat germ agglutinin as a specific probe for plasma membrane-containing fractions. The partly purified plasma membrane fraction had a density in sucrose of about 1.10 and was enriched in the activities of 5′-nucleotidase, alkaline phosphatase, sodium-potassium, and magnesium- dependent ATPases and adenylate cyclase. It contained only very low levels of acid phosphatase, cytochrome c oxidase, insulin, and RNA. Further purification was hampered by the relatively small amounts of fresh plasma membrane material that could be obtained from 16-24 rats in each experiment. When islets were prelabeled with radioactive fucose, the plasma membrane-enriched fraction contained radioactivity at a four- to fivefold higher specific activity than the whole islet homogenate. Sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis of plasma membrane-enriched fractions pooled from several experiments revealed a distinctive pattern of protein bands as compared with other less pure fractions. With respect to rapidity, apparent specificity, and easy reversibility of the labeling of the plasma membrane fraction, 125I-wheat germ agglutinin provides a highly useful tool for the detection of microgram quantities of plasma membrane components which should be applicable to many other systems as well.
(Less)
- author
- Lernmark, ÅKe LU ; Nathans, Anne and Steiner, Donald F.
- publishing date
- 1976-11-01
- type
- Contribution to journal
- publication status
- published
- in
- Journal of Cell Biology
- volume
- 71
- issue
- 2
- pages
- 606 - 623
- publisher
- Rockefeller University Press
- external identifiers
-
- scopus:0017188694
- pmid:791956
- ISSN
- 0021-9525
- DOI
- 10.1083/jcb.71.2.606
- language
- English
- LU publication?
- no
- id
- 637ea260-42de-4d62-b780-0d7eaed7dd2b
- date added to LUP
- 2019-09-18 12:12:13
- date last changed
- 2024-04-02 16:34:08
@article{637ea260-42de-4d62-b780-0d7eaed7dd2b, abstract = {{<p>Methods have been developed for the isolation on a semi-micro scale of a plasma membrane-enriched fraction from rat islets of Langerhans. An important feature of these experiments is the use of 125I-labeled wheat germ agglutinin as a specific probe for plasma membrane-containing fractions. The partly purified plasma membrane fraction had a density in sucrose of about 1.10 and was enriched in the activities of 5′-nucleotidase, alkaline phosphatase, sodium-potassium, and magnesium- dependent ATPases and adenylate cyclase. It contained only very low levels of acid phosphatase, cytochrome c oxidase, insulin, and RNA. Further purification was hampered by the relatively small amounts of fresh plasma membrane material that could be obtained from 16-24 rats in each experiment. When islets were prelabeled with radioactive fucose, the plasma membrane-enriched fraction contained radioactivity at a four- to fivefold higher specific activity than the whole islet homogenate. Sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis of plasma membrane-enriched fractions pooled from several experiments revealed a distinctive pattern of protein bands as compared with other less pure fractions. With respect to rapidity, apparent specificity, and easy reversibility of the labeling of the plasma membrane fraction, <sup>125</sup>I-wheat germ agglutinin provides a highly useful tool for the detection of microgram quantities of plasma membrane components which should be applicable to many other systems as well.</p>}}, author = {{Lernmark, ÅKe and Nathans, Anne and Steiner, Donald F.}}, issn = {{0021-9525}}, language = {{eng}}, month = {{11}}, number = {{2}}, pages = {{606--623}}, publisher = {{Rockefeller University Press}}, series = {{Journal of Cell Biology}}, title = {{Preparation and characterization of plasma membrane-enriched fractions from rat pancreatic islets}}, url = {{http://dx.doi.org/10.1083/jcb.71.2.606}}, doi = {{10.1083/jcb.71.2.606}}, volume = {{71}}, year = {{1976}}, }