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NanoSIP: NanoSIMS Applications for Microbial Biology

  • Jennifer Pett-Ridge
  • Peter K. Weber
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 881)

Abstract

Recent advances in high-resolution imaging secondary ion mass spectrometry (SIMS) (J Biol 5: 20, 2006) have made isotopic tracing at the single-cell level a standard technique for microbial ecology and systems biology; elemental and metal cofactor analyses are also showing significant promise. For example, with the NanoSIMS, metabolic activities of single microbial cells can be tracked by imaging natural isotopic/elemental composition or isotope distribution after stable isotope probing. When linked to molecular visualization methods, such as in situ hybridization and antibody labeling, these techniques enable in situ function to be linked to microbial identity and gene expression. We broadly call this combination of methods nanoSIP, for nanometer-scale stable isotope probing. Here we present the primary materials and methods used for nanoSIP, with an emphasis on key steps and potential pitfalls. Applications to pure cultures, cocultures, and complex communities are discussed.

Key words

NanoSIMS Isotope tracers Metal cofactors Single-cell biology Stable isotope probing Sample preparation SEM TEM FIB Fluorescence FISH Antibody labeling 

Notes

Acknowledgments

We wish to acknowledge Ian Hutcheon, for his many years of mentorship, advice, and leadership of the LLNL SIMS group. Christina Ramon plays a critical role in helping to prepare and organize many of the samples we have discussed. This work was funded in part by the DOE OBER Genomic Science program and LLNL Laboratory Directed Research and Development funding and performed under the auspices of the US Department of Energy at Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344. This document was prepared as an account of work sponsored by an agency of the United States government. Neither the United States government nor Lawrence Livermore National Security, LLC, nor any of their employees makes any warranty, expressed or implied, or assumes any legal liability or responsibility for the accuracy, completeness, or usefulness of any information, apparatus, product, or process disclosed, or represents that its use would not infringe privately owned rights. Reference herein to any specific commercial product, process, or service by trade name, trademark, manufacturer, or otherwise does not necessarily constitute or imply its endorsement, recommendation, or favoring by the United States government or Lawrence Livermore National Security, LLC. The views and opinions of authors expressed herein do not necessarily state or reflect those of the United States government or Lawrence Livermore National Security, LLC, and shall not be used for advertising or product endorsement purposes.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Chemical Sciences DivisionLawrence Livermore National LaboratoryLivermoreUSA

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