Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways

Maj Linea Vestergaard; Aashir Awan; Caroline Becker Warzecha; Søren Tvorup Christensen; Claus Yding Andersen

doi:10.1007/7651_2014_127

Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways

Maj Linea Vestergaard, Aashir Awan, Caroline Becker Warzecha, Søren Tvorup Christensen, Claus Yding Andersen

11 Citations (Scopus)

Abstract

This chapter describes the procedures for immunofluorescence microscopy (IFM) and quantitative PCR (qPCR) analyses of human embryonic stem cells (hESCs) grown specifically under feeder-free conditions. A detailed protocol is provided outlining the steps from initially growing the cells, passaging onto 16-well glass chambers, and continuing with the general IFM and qPCR anlysis. The techniques are illustrated with results on cellular localization of transcriptional factors and components of the Hedgehog, Wnt, PDGF, and TGFβ signaling pathways to primary cilia in stem cell maintenance and differentiation. Furthermore, a sample qPCR experiment is experimentally shown illustrating that these techniques can be important tools in answering basic questions about hESC biology.

Original language	English
Title of host publication	Human Embryonic Stem Cell Protocols
Volume	1307
Publisher	Springer Science+Business Media
Publication date	2015
Pages	123-140
ISBN (Print)	978-1-4939-2667-1, 978-1-4939-2668-8
DOIs	https://doi.org/10.1007/7651_2014_127
Publication status	Published - 2015

Series	Methods in molecular biology (Clifton, N.J.)
ISSN	1064-3745

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Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways. / Vestergaard, Maj Linea; Awan, Aashir; Warzecha, Caroline Becker et al.
Human Embryonic Stem Cell Protocols. Vol. 1307 Springer Science+Business Media, 2015. p. 123-140 (Methods in molecular biology (Clifton, N.J.)).

Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review

Vestergaard, ML, Awan, A, Warzecha, CB, Christensen, ST & Andersen, CY 2015, Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways. in Human Embryonic Stem Cell Protocols. vol. 1307, Springer Science+Business Media, Methods in molecular biology (Clifton, N.J.), pp. 123-140. https://doi.org/10.1007/7651_2014_127

Vestergaard ML, Awan A, Warzecha CB, Christensen ST , Andersen CY. Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways. In Human Embryonic Stem Cell Protocols. Vol. 1307. Springer Science+Business Media. 2015. p. 123-140. (Methods in molecular biology (Clifton, N.J.)). doi: 10.1007/7651_2014_127

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Immunofluorescence Microscopy and mRNA Analysis of Human Embryonic Stem Cells (hESCs) Including Primary Cilia Associated Signaling Pathways

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