Measuring Early Germ-Layer Specification Bias in Human Pluripotent Stem Cells

Alexander Keller; Nuša Krivec; Christina Markouli; Claudia Spits

doi:10.1007/978-1-0716-1979-7_5

Measuring Early Germ-Layer Specification Bias in Human Pluripotent Stem Cells

Methods Mol Biol. 2022:2429:57-72. doi: 10.1007/978-1-0716-1979-7_5.

Authors

Alexander Keller^#¹, Nuša Krivec^#¹, Christina Markouli¹, Claudia Spits²

Affiliations

¹ Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Jette, Belgium.
² Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Jette, Belgium. claudia.spits@vub.be.

^# Contributed equally.

PMID: 35507155
DOI: 10.1007/978-1-0716-1979-7_5

Abstract

Human pluripotent stem cells have a wide variety of potential applications, ranging from clinical translation to in vitro disease modeling. However, there is significant variation in the potential of individual cell lines to differentiate towards each of the three germ layers as a result of (epi)genetic background, culture conditions, and other factors. We describe here in detail a methodology to evaluate this bias using short directed differentiation towards neuroectoderm, mesendoderm, and definitive endoderm in combination with quantification by RT-qPCR and immunofluorescent stains.

Keywords: Definitive Endoderm; Differentiation Bias; Human Pluripotent Stem Cells; Mesendoderm; Neuroectoderm; Quantification of differentiation; Quantitative Immunofluorescent Stain; Quantitative Real-Time PCR.

MeSH terms

Cell Differentiation
Endoderm*
Germ Layers
Humans
Neural Plate
Pluripotent Stem Cells*