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Enrichment of single neurons and defined brain regions from human brain tissue samples for subsequent proteome analysis

Abstract

Brain function in normal aging and neurological diseases has long been a subject of interest. With current technology, it is possible to go beyond descriptive analyses to characterize brain cell populations at the molecular level. However, the brain comprises over 100 billion highly specialized cells, and it is a challenge to discriminate different cell groups for analyses. Isolating intact neurons is not feasible with traditional methods, such as tissue homogenization techniques. The advent of laser microdissection techniques promises to overcome previous limitations in the isolation of specific cells. Here, we provide a detailed protocol for isolating and analyzing neurons from postmortem human brain tissue samples. We describe a workflow for successfully freezing, sectioning and staining tissue for laser microdissection. This protocol was validated by mass spectrometric analysis. Isolated neurons can also be employed for western blotting or PCR. This protocol will enable further examinations of brain cell-specific molecular pathways and aid in elucidating distinct brain functions.

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Acknowledgments

This work was supported by WTZ Brasilien, a project of the BMBF, Germany (01DN14023), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPQ), Fundação de Amparo à Pesquisa do Estado de São Paulo (Fapesp), P.U.R.E. (Protein Unit for Research in Europe), a project of Nordrhein-Westfalen, a federal German state, and the HUPO Brain Proteome Project. The authors gratefully thank the Brazilian Brain Bank in São Paulo for providing tissues and Ulrich Sauer, Volker Wollscheid, Lukas Baran, Ulrike Weber and Gabrielle Friedemann from Carl Zeiss Microscopy GmbH, Munique, Germany, for technical support. Furthermore, we would like to thank Pascal C. Rauher for providing Fig. 1.

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There are no conflicts of interest to report.

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Correspondence to Katrin Marcus or Caroline May.

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M. Molina, S. Steinbach, R. E. P. Leite and C. May contributed equally.

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Molina, M., Steinbach, S., Park, Y.M. et al. Enrichment of single neurons and defined brain regions from human brain tissue samples for subsequent proteome analysis. J Neural Transm 122, 993–1005 (2015). https://doi.org/10.1007/s00702-015-1414-4

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Keywords

  • Neurons
  • Brain
  • Laser microdissection
  • Substantia nigra