A Multistep Workflow to Evaluate Newly Generated iPSCs and Their Ability to Generate Different Cell Types

Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of...

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Published inMethods and protocols Vol. 4; no. 3; p. 50
Main Authors Chen, Carol X.-Q., Abdian, Narges, Maussion, Gilles, Thomas, Rhalena A., Demirova, Iveta, Cai, Eddie, Tabatabaei, Mahdieh, Beitel, Lenore K., Karamchandani, Jason, Fon, Edward A., Durcan, Thomas M.
Format Journal Article
LanguageEnglish
Published Basel MDPI AG 19.07.2021
MDPI
Subjects
Alzheimer's disease
Cell differentiation
cortical neurons
Fibroblasts
Gene expression
Genomes
genomic integrity
Genomics
human-induced pluripotent stem cells
Inhibitory postsynaptic potentials
Morphology
neural progenitor cells
Neurogenesis
Pluripotency
quality control
Somatic cells
Stem cells
trilineage differentiation
Canada
United States > US
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Abstract Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of quality-control measures, promoting reproducibility across experiments and between labs. With this in mind, we established a multistep workflow to assess our newly generated iPSCs. Our workflow tests four benchmarks: cell growth, genomic stability, pluripotency, and the ability to form the three germline layers. We also outline a simple test for assessing cell growth and highlight the need to compare different growth media. Genomic integrity in the human iPSCs is analyzed by G-band karyotyping and a qPCR-based test for the detection of common karyotypic abnormalities. Finally, we confirm that the iPSC lines can differentiate into a given cell type, using a trilineage assay, and later confirm that each iPSC can be differentiated into one cell type of interest, with a focus on the generation of cortical neurons. Taken together, we present a multistep quality-control workflow to evaluate newly generated iPSCs and detail the findings on these lines as they are tested within the workflow.
AbstractList Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of quality-control measures, promoting reproducibility across experiments and between labs. With this in mind, we established a multistep workflow to assess our newly generated iPSCs. Our workflow tests four benchmarks: cell growth, genomic stability, pluripotency, and the ability to form the three germline layers. We also outline a simple test for assessing cell growth and highlight the need to compare different growth media. Genomic integrity in the human iPSCs is analyzed by G-band karyotyping and a qPCR-based test for the detection of common karyotypic abnormalities. Finally, we confirm that the iPSC lines can differentiate into a given cell type, using a trilineage assay, and later confirm that each iPSC can be differentiated into one cell type of interest, with a focus on the generation of cortical neurons. Taken together, we present a multistep quality-control workflow to evaluate newly generated iPSCs and detail the findings on these lines as they are tested within the workflow.
Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of quality-control measures, promoting reproducibility across experiments and between labs. With this in mind, we established a multistep workflow to assess our newly generated iPSCs. Our workflow tests four benchmarks: cell growth, genomic stability, pluripotency, and the ability to form the three germline layers. We also outline a simple test for assessing cell growth and highlight the need to compare different growth media. Genomic integrity in the human iPSCs is analyzed by G-band karyotyping and a qPCR-based test for the detection of common karyotypic abnormalities. Finally, we confirm that the iPSC lines can differentiate into a given cell type, using a trilineage assay, and later confirm that each iPSC can be differentiated into one cell type of interest, with a focus on the generation of cortical neurons. Taken together, we present a multistep quality-control workflow to evaluate newly generated iPSCs and detail the findings on these lines as they are tested within the workflow.Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of quality-control measures, promoting reproducibility across experiments and between labs. With this in mind, we established a multistep workflow to assess our newly generated iPSCs. Our workflow tests four benchmarks: cell growth, genomic stability, pluripotency, and the ability to form the three germline layers. We also outline a simple test for assessing cell growth and highlight the need to compare different growth media. Genomic integrity in the human iPSCs is analyzed by G-band karyotyping and a qPCR-based test for the detection of common karyotypic abnormalities. Finally, we confirm that the iPSC lines can differentiate into a given cell type, using a trilineage assay, and later confirm that each iPSC can be differentiated into one cell type of interest, with a focus on the generation of cortical neurons. Taken together, we present a multistep quality-control workflow to evaluate newly generated iPSCs and detail the findings on these lines as they are tested within the workflow.
Author Chen, Carol X.-Q.
Durcan, Thomas M.
Thomas, Rhalena A.
Beitel, Lenore K.
Karamchandani, Jason
Tabatabaei, Mahdieh
Fon, Edward A.
Maussion, Gilles
Cai, Eddie
Demirova, Iveta
Abdian, Narges
AuthorAffiliation 2 The Neuro’s Clinical Biological Imaging and Genetic Repository (C-BIG), McGill University, 3801 University Street, Montreal, QC H3A 2B4, Canada; ma.tabatabaei@mcgill.ca (M.T.); jason.karamchandani@mcgill.ca (J.K.)
1 The Neuro’s Early Drug Discovery Unit (EDDU), McGill University, 3801 University Street, Montreal, QC H3A 2B4, Canada; xiuqing.chen@mcgill.ca (C.X.-Q.C.); narges.abdian@mcgill.ca (N.A.); gilles.maussion@mcgill.ca (G.M.); rhalena.thomas@mcgill.ca (R.A.T.); iveta.demirova@mail.mcgill.ca (I.D.); eddie.cai@mail.mcgill.ca (E.C.); lenore.beitel@mcgill.ca (L.K.B.); ted.fon@mcgill.ca (E.A.F.)
AuthorAffiliation_xml – name: 2 The Neuro’s Clinical Biological Imaging and Genetic Repository (C-BIG), McGill University, 3801 University Street, Montreal, QC H3A 2B4, Canada; ma.tabatabaei@mcgill.ca (M.T.); jason.karamchandani@mcgill.ca (J.K.)
– name: 1 The Neuro’s Early Drug Discovery Unit (EDDU), McGill University, 3801 University Street, Montreal, QC H3A 2B4, Canada; xiuqing.chen@mcgill.ca (C.X.-Q.C.); narges.abdian@mcgill.ca (N.A.); gilles.maussion@mcgill.ca (G.M.); rhalena.thomas@mcgill.ca (R.A.T.); iveta.demirova@mail.mcgill.ca (I.D.); eddie.cai@mail.mcgill.ca (E.C.); lenore.beitel@mcgill.ca (L.K.B.); ted.fon@mcgill.ca (E.A.F.)
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Snippet Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of...
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SubjectTerms Alzheimer's disease
Cell differentiation
cortical neurons
Fibroblasts
Gene expression
Genomes
genomic integrity
Genomics
human-induced pluripotent stem cells
Inhibitory postsynaptic potentials
Morphology
neural progenitor cells
Neurogenesis
Pluripotency
quality control
Somatic cells
Stem cells
trilineage differentiation
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Title A Multistep Workflow to Evaluate Newly Generated iPSCs and Their Ability to Generate Different Cell Types
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