Cellular and Subcellular Immunohistochemical Localization and Quantification of Cadmium Ions in Wheat (Triticum aestivum)

• Published in: PloS one 2015-05, Vol.10 (5), p.e0123779-e0123779
• Main Authors: Gao, Wei, Nan, Tiegui, Tan, Guiyu, Zhao, Hongwei, Tan, Weiming, Meng, Fanyun, Li, Zhaohu, Li, Qing X, Wang, Baomin
• Format: Article
• Language: English
• Subjects: Agronomy; Arabidopsis; Bioengineering; Biotechnology; Brassica juncea; Cadmium; Cadmium - analysis; Cadmium - metabolism; Chelates; Confocal; Crops; Edetic acid; Ethylenediamine; Ethylenediaminetetraacetic acids; Fluorescence; Food; Gold; Heavy metals; Immunohistochemistry; Immunological tolerance; Ions; Localization; Measurement; Metals; Methods; Microscopy, Confocal; Microscopy, Electron, Transmission; Monoclonal antibodies; Physiology; Plant tissues; Proteins; Roots; Toxicity; Toxicity tolerance; Translocation; Triticum - chemistry; Triticum - metabolism; Triticum aestivum; Wheat; Zea mays
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0123779

The distribution of metallic ions in plant tissues is associated with their toxicity and is important for understanding mechanisms of toxicity tolerance. A quantitative histochemical method can help advance knowledge of cellular and subcellular localization and distribution of heavy metals in plant tissues. An immunohistochemical (IHC) imaging method for cadmium ions (Cd2+) was developed for the first time for the wheat Triticum aestivum grown in Cd2+-fortified soils. Also, 1-(4-Isothiocyanobenzyl)-ethylenediamine-N,N,N,N-tetraacetic acid (ITCB-EDTA) was used to chelate the mobile Cd2+. The ITCB-EDTA/Cd2+ complex was fixed with proteins in situ via the isothiocyano group. A new Cd2+-EDTA specific monoclonal antibody, 4F3B6D9A1, was used to locate the Cd2+-EDTA protein complex. After staining, the fluorescence intensities of sections of Cd2+-positive roots were compared with those of Cd2+-negative roots under a laser confocal scanning microscope, and the location of colloidal gold particles was determined with a transmission electron microscope. The results enable quantification of the Cd2+ content in plant tissues and illustrate Cd2+ translocation and cellular and subcellular responses of T. aestivum to Cd2+ stress. Compared to the conventional metal-S coprecipitation histochemical method, this new IHC method is quantitative, more specific and has less background interference. The subcellular location of Cd2+ was also confirmed with energy-dispersive X-ray microanalysis. The IHC method is suitable for locating and quantifying Cd2+ in plant tissues and can be extended to other heavy metallic ions.