Isolation of human bone marrow stromal cells from bone marrow biopsies for single-cell RNA sequencing

Hélène F.E. Gleitz; Inge A.M. Snoeren; Stijn N.R. Fuchs; Nils B. Leimkühler; Rebekka K. Schneider

doi:10.1016/j.xpro.2021.100538

Isolation of human bone marrow stromal cells from bone marrow biopsies for single-cell RNA sequencing

Hélène F.E. Gleitz^*, Inge A.M. Snoeren, Stijn N.R. Fuchs, Nils B. Leimkühler, Rebekka K. Schneider

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

6 Citations (Scopus)

Abstract

Bone marrow (BM) mesenchymal stromal cells play an important role in regulating stem cell quiescence and homeostasis; they are also key contributors to various hematological malignancies. However, human bone marrow stromal cells are difficult to isolate and prone to damage during isolation. This protocol describes a combination of mechanical and enzymatic isolation of BM stromal cells from human BM biopsies, followed by FACS sorting to separate stromal sub-populations including mesenchymal stromal cells, fibroblasts, and Schwann cells for single-cell RNA sequencing. For complete details on the use and execution of this protocol, please refer to Leimkühler et al. (2020).

Original language	English
Article number	100538
Journal	STAR Protocols
Volume	2
Issue number	2
DOIs	https://doi.org/10.1016/j.xpro.2021.100538
Publication status	Published - 18 Jun 2021

Bibliographical note

Funding Information:
H.F.E.G. was supported by the European Union’s Horizon 2020 research and innovation program under a Marie Curie-Sklodowska grant ( 707404 , LEaDing Fellow). N.B.L. was supported by the German Research Foundation, DFG ( BA 6349/1-1 ). R.K.S. is an Oncode Institute investigator and was supported by grants from the MPN Foundation ( 2017 MPNRF/LLS Award), a KWF Kankerbestrijding young investigator grant ( 11031/2017–1 , Bas Mulder Award; Dutch Cancer Foundation), and an ERC grant (deFIBER; ERC-StG 757339 ). This work was in part supported by grants of the Deutsche Forschungsgemeinschaft (DFG) (German Research Foundation) to R.K.S. ( SCHN1188/6-1 ) within the clinical research unit CRU344. R.K.S. is a member of the E:MED Consortia Fibromap funded by the German Ministry of Education and Science ( BMBF ). We thank the team of the Pathology Department at Erasmus Medical Center, Rotterdam.

Funding Information:
H.F.E.G. was supported by the European Union's Horizon 2020 research and innovation program under a Marie Curie-Sklodowska grant (707404, LEaDing Fellow). N.B.L. was supported by the German Research Foundation, DFG (BA 6349/1-1). R.K.S. is an Oncode Institute investigator and was supported by grants from the MPN Foundation (2017 MPNRF/LLS Award), a KWF Kankerbestrijding young investigator grant (11031/2017?1, Bas Mulder Award; Dutch Cancer Foundation), and an ERC grant (deFIBER; ERC-StG 757339). This work was in part supported by grants of the Deutsche Forschungsgemeinschaft (DFG) (German Research Foundation) to R.K.S. (SCHN1188/6-1) within the clinical research unit CRU344. R.K.S. is a member of the E:MED Consortia Fibromap funded by the German Ministry of Education and Science (BMBF). We thank the team of the Pathology Department at Erasmus Medical Center, Rotterdam. Conception and design, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; collection and assembly of data, H.F.E.G. I.A.M.S. S.N.R.F. and N.B.L.; data analysis and interpretation, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; manuscript writing, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; funding acquisition, R.K.S. The authors declare no competing interests.

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© 2021 The Authors

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10.1016/j.xpro.2021.100538

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title = "Isolation of human bone marrow stromal cells from bone marrow biopsies for single-cell RNA sequencing",

abstract = "Bone marrow (BM) mesenchymal stromal cells play an important role in regulating stem cell quiescence and homeostasis; they are also key contributors to various hematological malignancies. However, human bone marrow stromal cells are difficult to isolate and prone to damage during isolation. This protocol describes a combination of mechanical and enzymatic isolation of BM stromal cells from human BM biopsies, followed by FACS sorting to separate stromal sub-populations including mesenchymal stromal cells, fibroblasts, and Schwann cells for single-cell RNA sequencing. For complete details on the use and execution of this protocol, please refer to Leimk{\"u}hler et al. (2020).",

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note = "Funding Information: H.F.E.G. was supported by the European Union{\textquoteright}s Horizon 2020 research and innovation program under a Marie Curie-Sklodowska grant ( 707404 , LEaDing Fellow). N.B.L. was supported by the German Research Foundation, DFG ( BA 6349/1-1 ). R.K.S. is an Oncode Institute investigator and was supported by grants from the MPN Foundation ( 2017 MPNRF/LLS Award), a KWF Kankerbestrijding young investigator grant ( 11031/2017–1 , Bas Mulder Award; Dutch Cancer Foundation), and an ERC grant (deFIBER; ERC-StG 757339 ). This work was in part supported by grants of the Deutsche Forschungsgemeinschaft (DFG) (German Research Foundation) to R.K.S. ( SCHN1188/6-1 ) within the clinical research unit CRU344. R.K.S. is a member of the E:MED Consortia Fibromap funded by the German Ministry of Education and Science ( BMBF ). We thank the team of the Pathology Department at Erasmus Medical Center, Rotterdam. Funding Information: H.F.E.G. was supported by the European Union's Horizon 2020 research and innovation program under a Marie Curie-Sklodowska grant (707404, LEaDing Fellow). N.B.L. was supported by the German Research Foundation, DFG (BA 6349/1-1). R.K.S. is an Oncode Institute investigator and was supported by grants from the MPN Foundation (2017 MPNRF/LLS Award), a KWF Kankerbestrijding young investigator grant (11031/2017?1, Bas Mulder Award; Dutch Cancer Foundation), and an ERC grant (deFIBER; ERC-StG 757339). This work was in part supported by grants of the Deutsche Forschungsgemeinschaft (DFG) (German Research Foundation) to R.K.S. (SCHN1188/6-1) within the clinical research unit CRU344. R.K.S. is a member of the E:MED Consortia Fibromap funded by the German Ministry of Education and Science (BMBF). We thank the team of the Pathology Department at Erasmus Medical Center, Rotterdam. Conception and design, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; collection and assembly of data, H.F.E.G. I.A.M.S. S.N.R.F. and N.B.L.; data analysis and interpretation, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; manuscript writing, H.F.E.G. I.A.M.S. S.N.R.F. R.K.S. and N.B.L.; funding acquisition, R.K.S. The authors declare no competing interests. Publisher Copyright: {\textcopyright} 2021 The Authors",

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AU - Snoeren, Inge A.M.

AU - Fuchs, Stijn N.R.

AU - Leimkühler, Nils B.

AU - Schneider, Rebekka K.

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N2 - Bone marrow (BM) mesenchymal stromal cells play an important role in regulating stem cell quiescence and homeostasis; they are also key contributors to various hematological malignancies. However, human bone marrow stromal cells are difficult to isolate and prone to damage during isolation. This protocol describes a combination of mechanical and enzymatic isolation of BM stromal cells from human BM biopsies, followed by FACS sorting to separate stromal sub-populations including mesenchymal stromal cells, fibroblasts, and Schwann cells for single-cell RNA sequencing. For complete details on the use and execution of this protocol, please refer to Leimkühler et al. (2020).

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Isolation of human bone marrow stromal cells from bone marrow biopsies for single-cell RNA sequencing

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