A trichloroacetic acid-acetone method greatly reduces infrared autofluorescence of protein extracts from plant tissue

Immunoblotting is used to determine many important characteristics of proteins. After electrophoretic separation, proteins are transferred to a membrane and reacted with a specific antibody. The antibody-protein complex is then visualized by radiographic, chromogenic, chemiluminescent, or more recen...

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Bibliographic Details
Published in:Plant molecular biology reporter 2005-12, Vol.23 (4), p.405-409
Main Authors: Shultz, Randall W., Settlage, Sharon B., Hanley-Bowdoin, Linda, Thompson, William F.
Format: Article
Language:English
Subjects:
Acetone
Antigens
Chemiluminescence
Fluorescence
Immunoblotting
Immunogenicity
Infrared analysis
Pigments
Plant extracts
Plant tissues
Proteins
Trichloroacetic acid
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Summary:Immunoblotting is used to determine many important characteristics of proteins. After electrophoretic separation, proteins are transferred to a membrane and reacted with a specific antibody. The antibody-protein complex is then visualized by radiographic, chromogenic, chemiluminescent, or more recently described fluorescence detection methods. Fluorescence-based detection offers some advantages over other approaches, including increased sensitivity, improved quantifiable range, and the ability to detect multiple antigens on the same blot. However, this technique is unavailable for analysis of green plant tissues by standard extraction methods because of contaminating autofluorescent pigments. We have compared 3 methods for extracting protein from plant tissue for use with infrared fluorescence-based immunoblot analysis. We report a trichloroacetic acid-acetone method that effectively eliminates autofluorescence while retaining the immunogenicity of a target protein.
ISSN:0735-9640
1572-9818
DOI:10.1007/BF02788888