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

Abstract

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. © 2005 International Society for Plant Molecular Biology.