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A laboratory exercise illustrating the sensitivity and specificity of Western blot analysis.
MedLine Citation:
PMID:  21774057     Owner:  NLM     Status:  In-Data-Review    
Abstract/OtherAbstract:
Western blot analysis, commonly known as "Western blotting," is a standard tool in every laboratory where proteins are analyzed. It involves the separation of polypeptides in polyacrylamide gels followed by the electrophoretic transfer of the separated polypeptides onto a nitrocellulose or polyvinylidene fluoride membrane. A replica of the separated polypeptides from the gel is created on the membrane, which is then probed with antibodies or other ligands to identify specific polypeptide(s). Here, we report an undergraduate laboratory exercise involving Western blotting. During the 3-week laboratory exercise, students investigated the likelihood of the presence of a serum albumin in fruit fly (Drosophila melanogaster) homogenate, bovine calf serum (BCS), and fetal bovine serum (FBS). Students isolate proteins from fruit fly larvae and perform sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE), electrophoretic blotting, and immunoassay comparing those proteins with BCS and FBS proteins. Their results indicate that serum albumin is present in BCS and FBS but is absent in fruit flies. In the process, the specificity and sensitivity of Western blot analysis is demonstrated. The laboratory exercise can be easily incorporated into any college-level biochemistry or molecular techniques laboratory. The procedure used can be easily adapted to study other proteins. Biochemistry and Molecular Biology Education Vol. 39, No. 4, pp. 291-297, 2011.
Authors:
Ming-Mei Chang; Janice Lovett
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Biochemistry and molecular biology education : a bimonthly publication of the International Union of Biochemistry and Molecular Biology     Volume:  39     ISSN:  1539-3429     ISO Abbreviation:  Biochem Mol Biol Educ     Publication Date:  2011 Jul 
Date Detail:
Created Date:  2011-07-20     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  100970605     Medline TA:  Biochem Mol Biol Educ     Country:  United States    
Other Details:
Languages:  eng     Pagination:  291-7     Citation Subset:  IM    
Copyright Information:
Copyright © 2011 Wiley Periodicals, Inc.
Affiliation:
Department of Biology, State University of New York at Geneseo, Geneseo, New York 14454. chang@geneseo.edu.
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