Flow cytometric sorting of neuronal and glial nuclei from central nervous system tissue

Seiji Okada, Hirokazu Saiwai, Hiromi Kumamaru, Kensuke Kubota, Akihito Harada, Masahiro Yamaguchi, Yukihide Iwamoto, Yasuyuki Ohkawa

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Due to the complex cellular heterogeneity of the central nervous system (CNS), it is relatively difficult to reliably obtain molecular descriptions with cell-type specificity. In particular, comparative analysis of epigenetic regulation or molecular profiles is hampered by the lack of adequate methodology for selective purification of defined cell populations from CNS tissue. Here, we developed a direct purification strategy of neural nuclei from CNS tissue based on fluorescence-activated cell sorting (FACS). We successfully fractionated nuclei from complex tissues such as brain, spinal cord, liver, kidney, and skeletal muscle extruded mechanically or chemically, and fractionated nuclei were structurally maintained and contained nucleoproteins and nuclear DNA/RNA. We collected sufficient numbers of nuclei from neurons and oligodendrocytes using FACS with immunolabeling for nucleoproteins or from genetically labeled transgenic mice. In addition, the use of Fab fragments isolated from papain antibody digests, which effectively enriched the specialized cell populations, significantly enhanced the immunolabeling efficacy. This methodology can be applied to a wide variety of heterogeneous tissues and is crucial for understanding the cell-specific information about chromatin dynamics, nucleoproteins, protein-DNA/RNA interactions, and transcriptomes retained in the nucleus, such as non-coding RNAs.

Original languageEnglish
Pages (from-to)552-558
Number of pages7
JournalJournal of cellular physiology
Volume226
Issue number2
DOIs
Publication statusPublished - Feb 1 2011

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Nerve Tissue
Neurology
Sorting
Neuroglia
Nucleoproteins
Central Nervous System
Tissue
Cells
Purification
Flow Cytometry
Fluorescence
RNA
Nuclear RNA
Untranslated RNA
Immunoglobulin Fab Fragments
Papain
DNA
Oligodendroglia
Transcriptome
Epigenomics

All Science Journal Classification (ASJC) codes

  • Physiology
  • Clinical Biochemistry
  • Cell Biology

Cite this

Flow cytometric sorting of neuronal and glial nuclei from central nervous system tissue. / Okada, Seiji; Saiwai, Hirokazu; Kumamaru, Hiromi; Kubota, Kensuke; Harada, Akihito; Yamaguchi, Masahiro; Iwamoto, Yukihide; Ohkawa, Yasuyuki.

In: Journal of cellular physiology, Vol. 226, No. 2, 01.02.2011, p. 552-558.

Research output: Contribution to journalArticle

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