Gold-ISH: A nano-size gold particle-based phylogenetic identification compatible with NanoSIMS

Kengo Kubota, Yuki Morono, Motoo Ito, Takeshi Terada, Shogo Itezono, Hideki Harada, Fumio Inagaki

Research output: Contribution to journalArticlepeer-review

9 Citations (Scopus)

Abstract

The linkage of microbial phylogenetic and metabolic analyses by combining ion imaging analysis with nano-scale secondary ion mass spectrometry (NanoSIMS) has become a powerful means of exploring the metabolic functions of environmental microorganisms. Phylogenetic identification using NanoSIMS typically involves probing by horseradish peroxidase-mediated deposition of halogenated fluorescent tyramides, which permits highly sensitive detection of specific microbial cells. However, the methods require permeabilization of target microbial cells and inactivation of endogenous peroxidase activity, and the use of halogens as the target atom is limited because of heavy background signals due to the presence of halogenated minerals in soil and sediment samples. Here, we present "Gold-ISH," a non-halogen phylogenetic probing method in which oligonucleotide probes are directly labeled with Undecagold, an ultra-small gold nanoparticle. Undecagold-labeled probes were generated using a thiol-maleimide chemical coupling reaction and they were purified by polyacrylamide gel electrophoresis. The method was optimized with a mixture of axenic 13C-labeled Escherichia coli and Methanococcus maripaludis cells and applied to investigate sulfate-reducing bacteria in an anaerobic sludge sample. Clear gold-derived target signals were detected in microbial cells using NanoSIMS ion imaging. It was concluded that Gold-ISH can be a useful approach for metabolic studies of naturally occurring microbial ecosystems using NanoSIMS.

Original languageEnglish
Pages (from-to)261-266
Number of pages6
JournalSystematic and Applied Microbiology
Volume37
Issue number4
DOIs
Publication statusPublished - 2014 Jun

Keywords

  • Gold-ISH
  • NanoSIMS ion microprobe
  • Undecagold

ASJC Scopus subject areas

  • Microbiology
  • Ecology, Evolution, Behavior and Systematics
  • Applied Microbiology and Biotechnology

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