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Updated: May 30, 2026

Preparation of Chloroplast Sub-compartments from Arabidopsis for the Analysis of Protein Localization by Immunoblotting or Proteomics
Published on: October 19, 2018
Immunofluorescence microscopy for localization of Arabidopsis chloroplast proteins
Stanislav Vitha1, Katherine W Osteryoung
1Microscopy and Imaging Center, Texas A&M University, College Station, TX, USA.
Abstract:
Immunofluorescence microscopy reveals localization of proteins in cells and tissues by means of highly specific, fluorescently labeled antibodies. This technique is an important complement to localization methods that use genetically encoded fluorescent tags. This chapter describes the five stages of immunofluorescence localization of proteins in plant chloroplasts in sectioned leaf tissue: (1) fixation, (2) tissue embedding and sectioning, (3) treatment of sections prior to immunolabeling, (4) immunostaining, and (5) fluorescence microscopy and image capture. Protocols for both cryosectioning and sectioning of low-melting-point wax-embedded samples are described. Immunofluorescence localization in chloroplasts is complicated by their intense autofluorescence background. Measures to suppress nonspecific background staining, confirm specificity of the fluorescence signal, and optimize imaging conditions are described.

