---
_id: '14683'
abstract:
- lang: eng
  text: "Mosaic analysis with double markers (MADM) technology enables the generation
    of genetic mosaic tissue in mice and high-resolution phenotyping at the individual
    cell level. Here, we present a protocol for isolating MADM-labeled cells with
    high yield for downstream molecular analyses using fluorescence-activated cell
    sorting (FACS). We describe steps for generating MADM-labeled mice, perfusion,
    single-cell suspension, and debris removal. We then detail procedures for cell
    sorting by FACS and downstream analysis. This protocol is suitable for embryonic
    to adult mice.\r\nFor complete details on the use and execution of this protocol,
    please refer to Contreras et al. (2021).1"
acknowledged_ssus:
- _id: Bio
- _id: PreCl
acknowledgement: This research was supported by the Scientific Service Units (SSU)
  at IST Austria through resources provided by the Imaging & Optics Facility (IOF)
  and Preclinical Facilities (PCF). N.A. received support from FWF Firnberg-Programme
  (T 1031). G.C. received support from the European Union’s Horizon 2020 research
  and innovation programme under the Marie Skłodowska-Curie grant agreement no. 754411
  as an ISTplus postdoctoral fellow. This work was also supported by IST Austria institutional
  funds, FWF SFB F78 to S.H., and the European Research Council (ERC) under the European
  Union’s Horizon 2020 research and innovation programme (grant agreement no. 725780
  LinPro) to S.H.
article_number: '102771'
article_processing_charge: No
article_type: review
author:
- first_name: Nicole
  full_name: Amberg, Nicole
  id: 4CD6AAC6-F248-11E8-B48F-1D18A9856A87
  last_name: Amberg
  orcid: 0000-0002-3183-8207
- first_name: Giselle T
  full_name: Cheung, Giselle T
  id: 471195F6-F248-11E8-B48F-1D18A9856A87
  last_name: Cheung
  orcid: 0000-0001-8457-2572
- first_name: Simon
  full_name: Hippenmeyer, Simon
  id: 37B36620-F248-11E8-B48F-1D18A9856A87
  last_name: Hippenmeyer
  orcid: 0000-0003-2279-1061
citation:
  ama: Amberg N, Cheung GT, Hippenmeyer S. Protocol for sorting cells from mouse brains
    labeled with mosaic analysis with double markers by flow cytometry. <i>STAR Protocols</i>.
    2023;5(1). doi:<a href="https://doi.org/10.1016/j.xpro.2023.102771">10.1016/j.xpro.2023.102771</a>
  apa: Amberg, N., Cheung, G. T., &#38; Hippenmeyer, S. (2023). Protocol for sorting
    cells from mouse brains labeled with mosaic analysis with double markers by flow
    cytometry. <i>STAR Protocols</i>. Elsevier. <a href="https://doi.org/10.1016/j.xpro.2023.102771">https://doi.org/10.1016/j.xpro.2023.102771</a>
  chicago: Amberg, Nicole, Giselle T Cheung, and Simon Hippenmeyer. “Protocol for
    Sorting Cells from Mouse Brains Labeled with Mosaic Analysis with Double Markers
    by Flow Cytometry.” <i>STAR Protocols</i>. Elsevier, 2023. <a href="https://doi.org/10.1016/j.xpro.2023.102771">https://doi.org/10.1016/j.xpro.2023.102771</a>.
  ieee: N. Amberg, G. T. Cheung, and S. Hippenmeyer, “Protocol for sorting cells from
    mouse brains labeled with mosaic analysis with double markers by flow cytometry,”
    <i>STAR Protocols</i>, vol. 5, no. 1. Elsevier, 2023.
  ista: Amberg N, Cheung GT, Hippenmeyer S. 2023. Protocol for sorting cells from
    mouse brains labeled with mosaic analysis with double markers by flow cytometry.
    STAR Protocols. 5(1), 102771.
  mla: Amberg, Nicole, et al. “Protocol for Sorting Cells from Mouse Brains Labeled
    with Mosaic Analysis with Double Markers by Flow Cytometry.” <i>STAR Protocols</i>,
    vol. 5, no. 1, 102771, Elsevier, 2023, doi:<a href="https://doi.org/10.1016/j.xpro.2023.102771">10.1016/j.xpro.2023.102771</a>.
  short: N. Amberg, G.T. Cheung, S. Hippenmeyer, STAR Protocols 5 (2023).
date_created: 2023-12-13T11:48:05Z
date_published: 2023-12-08T00:00:00Z
date_updated: 2023-12-18T08:06:14Z
day: '08'
ddc:
- '570'
department:
- _id: SiHi
doi: 10.1016/j.xpro.2023.102771
ec_funded: 1
external_id:
  pmid:
  - '38070137'
intvolume: '         5'
issue: '1'
keyword:
- General Immunology and Microbiology
- General Biochemistry
- Genetics and Molecular Biology
- General Neuroscience
language:
- iso: eng
main_file_link:
- open_access: '1'
  url: https://doi.org/10.1016/j.xpro.2023.102771
month: '12'
oa: 1
oa_version: Submitted Version
pmid: 1
project:
- _id: 268F8446-B435-11E9-9278-68D0E5697425
  call_identifier: FWF
  grant_number: T0101031
  name: Role of Eed in neural stem cell lineage progression
- _id: 260C2330-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '754411'
  name: ISTplus - Postdoctoral Fellowships
- _id: 059F6AB4-7A3F-11EA-A408-12923DDC885E
  grant_number: F07805
  name: Molecular Mechanisms of Neural Stem Cell Lineage Progression
- _id: 260018B0-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '725780'
  name: Principles of Neural Stem Cell Lineage Progression in Cerebral Cortex Development
publication: STAR Protocols
publication_identifier:
  issn:
  - 2666-1667
publication_status: epub_ahead
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Protocol for sorting cells from mouse brains labeled with mosaic analysis with
  double markers by flow cytometry
tmp:
  image: /images/cc_by.png
  legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
  name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
  short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 5
year: '2023'
...
---
_id: '12117'
abstract:
- lang: eng
  text: "To understand how potential gene manipulations affect in vitro microglia,
    we provide a set of short protocols to evaluate microglia identity and function.
    We detail steps for immunostaining to determine microglia identity. We describe
    three functional assays for microglia: phagocytosis, calcium response following
    ATP stimulation, and cytokine expression upon inflammatory stimuli. We apply these
    protocols to human induced-pluripotent-stem-cell (hiPSC)-derived microglia, but
    they can be also applied to other in vitro microglial models including primary
    mouse microglia.\r\nFor complete details on the use and execution of this protocol,
    please refer to Bartalska et al. (2022).1"
acknowledged_ssus:
- _id: Bio
acknowledgement: This project has received funding from the European Research Council
  (ERC) under the European Union’s Horizon 2020 research and innovation program (grant
  No. 715571 to S.S.) and from the Gesellschaft für Forschungsförderung Niederösterreich
  (grant No. Sc19-017 to V.H.). We thank Rouven Schulz and Alessandro Venturino for
  their insights into functional assays and data analysis, Verena Seiboth for insights
  into necessary institutional permission, and ISTA imaging & optics facility (IOF)
  especially Bernhard Hochreiter for their support.
article_number: '101866'
article_processing_charge: No
article_type: letter_note
author:
- first_name: Verena
  full_name: Hübschmann, Verena
  id: 32B7C918-F248-11E8-B48F-1D18A9856A87
  last_name: Hübschmann
- first_name: Medina
  full_name: Korkut, Medina
  id: 4B51CE74-F248-11E8-B48F-1D18A9856A87
  last_name: Korkut
  orcid: 0000-0003-4309-2251
- first_name: Sandra
  full_name: Siegert, Sandra
  id: 36ACD32E-F248-11E8-B48F-1D18A9856A87
  last_name: Siegert
  orcid: 0000-0001-8635-0877
citation:
  ama: Hübschmann V, Korkut M, Siegert S. Assessing human iPSC-derived microglia identity
    and function by immunostaining, phagocytosis, calcium activity, and inflammation
    assay. <i>STAR Protocols</i>. 2022;3(4). doi:<a href="https://doi.org/10.1016/j.xpro.2022.101866">10.1016/j.xpro.2022.101866</a>
  apa: Hübschmann, V., Korkut, M., &#38; Siegert, S. (2022). Assessing human iPSC-derived
    microglia identity and function by immunostaining, phagocytosis, calcium activity,
    and inflammation assay. <i>STAR Protocols</i>. Elsevier. <a href="https://doi.org/10.1016/j.xpro.2022.101866">https://doi.org/10.1016/j.xpro.2022.101866</a>
  chicago: Hübschmann, Verena, Medina Korkut, and Sandra Siegert. “Assessing Human
    IPSC-Derived Microglia Identity and Function by Immunostaining, Phagocytosis,
    Calcium Activity, and Inflammation Assay.” <i>STAR Protocols</i>. Elsevier, 2022.
    <a href="https://doi.org/10.1016/j.xpro.2022.101866">https://doi.org/10.1016/j.xpro.2022.101866</a>.
  ieee: V. Hübschmann, M. Korkut, and S. Siegert, “Assessing human iPSC-derived microglia
    identity and function by immunostaining, phagocytosis, calcium activity, and inflammation
    assay,” <i>STAR Protocols</i>, vol. 3, no. 4. Elsevier, 2022.
  ista: Hübschmann V, Korkut M, Siegert S. 2022. Assessing human iPSC-derived microglia
    identity and function by immunostaining, phagocytosis, calcium activity, and inflammation
    assay. STAR Protocols. 3(4), 101866.
  mla: Hübschmann, Verena, et al. “Assessing Human IPSC-Derived Microglia Identity
    and Function by Immunostaining, Phagocytosis, Calcium Activity, and Inflammation
    Assay.” <i>STAR Protocols</i>, vol. 3, no. 4, 101866, Elsevier, 2022, doi:<a href="https://doi.org/10.1016/j.xpro.2022.101866">10.1016/j.xpro.2022.101866</a>.
  short: V. Hübschmann, M. Korkut, S. Siegert, STAR Protocols 3 (2022).
date_created: 2023-01-12T11:56:38Z
date_published: 2022-12-16T00:00:00Z
date_updated: 2023-11-02T12:21:32Z
day: '16'
ddc:
- '570'
department:
- _id: SaSi
- _id: GradSch
doi: 10.1016/j.xpro.2022.101866
ec_funded: 1
file:
- access_level: open_access
  checksum: 3c71b8a60633d42c2f77c49025d5559b
  content_type: application/pdf
  creator: dernst
  date_created: 2023-01-23T09:50:51Z
  date_updated: 2023-01-23T09:50:51Z
  file_id: '12340'
  file_name: 2022_STARProtocols_Huebschmann.pdf
  file_size: 6251945
  relation: main_file
  success: 1
file_date_updated: 2023-01-23T09:50:51Z
has_accepted_license: '1'
intvolume: '         3'
issue: '4'
keyword:
- General Immunology and Microbiology
- General Biochemistry
- Genetics and Molecular Biology
- General Neuroscience
language:
- iso: eng
month: '12'
oa: 1
oa_version: Published Version
project:
- _id: 25D4A630-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '715571'
  name: Microglia action towards neuronal circuit formation and function in health
    and disease
- _id: 9B99D380-BA93-11EA-9121-9846C619BF3A
  grant_number: SC19-017
  name: How human microglia shape developing neurons during health and inflammation
publication: STAR Protocols
publication_identifier:
  issn:
  - 2666-1667
publication_status: published
publisher: Elsevier
quality_controlled: '1'
related_material:
  record:
  - id: '11478'
    relation: other
    status: public
scopus_import: '1'
status: public
title: Assessing human iPSC-derived microglia identity and function by immunostaining,
  phagocytosis, calcium activity, and inflammation assay
tmp:
  image: /images/cc_by_nc_nd.png
  legal_code_url: https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode
  name: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
    (CC BY-NC-ND 4.0)
  short: CC BY-NC-ND (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 3
year: '2022'
...
---
_id: '8978'
abstract:
- lang: eng
  text: "Mosaic analysis with double markers (MADM) technology enables concomitant
    fluorescent cell labeling and induction of uniparental chromosome disomy (UPD)
    with single-cell resolution. In UPD, imprinted genes are either overexpressed
    2-fold or are not expressed. Here, the MADM platform is utilized to probe imprinting
    phenotypes at the transcriptional level. This protocol highlights major steps
    for the generation and isolation of projection neurons and astrocytes with MADM-induced
    UPD from mouse cerebral cortex for downstream single-cell and low-input sample
    RNA-sequencing experiments.\r\n\r\nFor complete details on the use and execution
    of this protocol, please refer to Laukoter et al. (2020b)."
acknowledged_ssus:
- _id: Bio
- _id: PreCl
acknowledgement: This research was supported by the Scientific Service Units (SSU)
  at IST Austria through resources provided by the Bioimaging (BIF) and Preclinical
  Facilities (PCF). N.A received support from the FWF Firnberg-Programm (T 1031).
  This work was also supported by IST Austria institutional funds; FWF SFB F78 to
  S.H.; NÖ Forschung und Bildung n[f+b] life science call grant (C13-002) to S.H.;
  the People Programme (Marie Curie Actions) of the European Union’s Seventh Framework
  Programme (FP7/2007-2013) under REA grant agreement no. 618444 to S.H.; and the
  European Research Council (ERC) under the European Union’s Horizon 2020 research
  and innovation programme (grant agreement no. 725780 LinPro) to S.H.
article_number: '100215'
article_processing_charge: No
article_type: original
author:
- first_name: Susanne
  full_name: Laukoter, Susanne
  id: 2D6B7A9A-F248-11E8-B48F-1D18A9856A87
  last_name: Laukoter
- first_name: Nicole
  full_name: Amberg, Nicole
  id: 4CD6AAC6-F248-11E8-B48F-1D18A9856A87
  last_name: Amberg
  orcid: 0000-0002-3183-8207
- first_name: Florian
  full_name: Pauler, Florian
  id: 48EA0138-F248-11E8-B48F-1D18A9856A87
  last_name: Pauler
- first_name: Simon
  full_name: Hippenmeyer, Simon
  id: 37B36620-F248-11E8-B48F-1D18A9856A87
  last_name: Hippenmeyer
  orcid: 0000-0003-2279-1061
citation:
  ama: Laukoter S, Amberg N, Pauler F, Hippenmeyer S. Generation and isolation of
    single cells from mouse brain with mosaic analysis with double markers-induced
    uniparental chromosome disomy. <i>STAR Protocols</i>. 2020;1(3). doi:<a href="https://doi.org/10.1016/j.xpro.2020.100215">10.1016/j.xpro.2020.100215</a>
  apa: Laukoter, S., Amberg, N., Pauler, F., &#38; Hippenmeyer, S. (2020). Generation
    and isolation of single cells from mouse brain with mosaic analysis with double
    markers-induced uniparental chromosome disomy. <i>STAR Protocols</i>. Elsevier.
    <a href="https://doi.org/10.1016/j.xpro.2020.100215">https://doi.org/10.1016/j.xpro.2020.100215</a>
  chicago: Laukoter, Susanne, Nicole Amberg, Florian Pauler, and Simon Hippenmeyer.
    “Generation and Isolation of Single Cells from Mouse Brain with Mosaic Analysis
    with Double Markers-Induced Uniparental Chromosome Disomy.” <i>STAR Protocols</i>.
    Elsevier, 2020. <a href="https://doi.org/10.1016/j.xpro.2020.100215">https://doi.org/10.1016/j.xpro.2020.100215</a>.
  ieee: S. Laukoter, N. Amberg, F. Pauler, and S. Hippenmeyer, “Generation and isolation
    of single cells from mouse brain with mosaic analysis with double markers-induced
    uniparental chromosome disomy,” <i>STAR Protocols</i>, vol. 1, no. 3. Elsevier,
    2020.
  ista: Laukoter S, Amberg N, Pauler F, Hippenmeyer S. 2020. Generation and isolation
    of single cells from mouse brain with mosaic analysis with double markers-induced
    uniparental chromosome disomy. STAR Protocols. 1(3), 100215.
  mla: Laukoter, Susanne, et al. “Generation and Isolation of Single Cells from Mouse
    Brain with Mosaic Analysis with Double Markers-Induced Uniparental Chromosome
    Disomy.” <i>STAR Protocols</i>, vol. 1, no. 3, 100215, Elsevier, 2020, doi:<a
    href="https://doi.org/10.1016/j.xpro.2020.100215">10.1016/j.xpro.2020.100215</a>.
  short: S. Laukoter, N. Amberg, F. Pauler, S. Hippenmeyer, STAR Protocols 1 (2020).
date_created: 2020-12-30T10:17:07Z
date_published: 2020-12-18T00:00:00Z
date_updated: 2021-01-12T08:21:36Z
day: '18'
ddc:
- '570'
department:
- _id: SiHi
doi: 10.1016/j.xpro.2020.100215
ec_funded: 1
external_id:
  pmid:
  - '33377108'
file:
- access_level: open_access
  checksum: f1e9a433e9cb0f41f7b6df6b76db1f6e
  content_type: application/pdf
  creator: dernst
  date_created: 2021-01-07T15:57:27Z
  date_updated: 2021-01-07T15:57:27Z
  file_id: '8996'
  file_name: 2020_STARProtocols_Laukoter.pdf
  file_size: 4031449
  relation: main_file
  success: 1
file_date_updated: 2021-01-07T15:57:27Z
has_accepted_license: '1'
intvolume: '         1'
issue: '3'
language:
- iso: eng
month: '12'
oa: 1
oa_version: Published Version
pmid: 1
project:
- _id: 268F8446-B435-11E9-9278-68D0E5697425
  call_identifier: FWF
  grant_number: T0101031
  name: Role of Eed in neural stem cell lineage progression
- _id: 059F6AB4-7A3F-11EA-A408-12923DDC885E
  grant_number: F07805
  name: Molecular Mechanisms of Neural Stem Cell Lineage Progression
- _id: 25D92700-B435-11E9-9278-68D0E5697425
  grant_number: LS13-002
  name: Mapping Cell-Type Specificity of the Genomic Imprintome in the Brain
- _id: 25D61E48-B435-11E9-9278-68D0E5697425
  call_identifier: FP7
  grant_number: '618444'
  name: Molecular Mechanisms of Cerebral Cortex Development
- _id: 260018B0-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '725780'
  name: Principles of Neural Stem Cell Lineage Progression in Cerebral Cortex Development
publication: STAR Protocols
publication_identifier:
  issn:
  - 2666-1667
publication_status: published
publisher: Elsevier
quality_controlled: '1'
status: public
title: Generation and isolation of single cells from mouse brain with mosaic analysis
  with double markers-induced uniparental chromosome disomy
tmp:
  image: /images/cc_by_nc_nd.png
  legal_code_url: https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode
  name: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
    (CC BY-NC-ND 4.0)
  short: CC BY-NC-ND (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 1
year: '2020'
...