Generation of Monoclonal Antibodies Specific for Cell Surface Molecules Expressed on Early Mouse Endoderm
Open Access
- 11 June 2009
- journal article
- research article
- Published by Oxford University Press (OUP) in The International Journal of Cell Cloning
- Vol. 27 (9), 2103-2113
- https://doi.org/10.1002/stem.147
Abstract
The development of functional cell populations such as hepatocytes and pancreatic β cells from embryonic stem cell (ESC) is dependent on the efficient induction of definitive endoderm early in the differentiation process. To monitor definitive endoderm formation in mouse ESC differentiation cultures in a quantitative fashion, we generated a reporter cell line that expresses human CD25 from the Foxa3 locus and human CD4 from the Foxa2 locus. Induction of these reporter ESCs with high concentrations of activin A led to the development of a CD25-Foxa3+CD4-Foxa2+ population within 4-5 days of culture. Isolation and characterization of this population showed that it consists predominantly of definitive endoderm that is able to undergo hepatic specification under the appropriate conditions. To develop reagents that can be used for studies on endoderm development from unmanipulated ESCs, from induced pluripotent stem cells, and from the mouse embryo, we generated monoclonal antibodies against the CD25-Foxa3+CD4-Foxa2+ population. With this approach, we identified two antibodies that react specifically with endoderm from ESC cultures and from the early embryo. The specificity of these antibodies enables one to quantitatively monitor endoderm development in ESC differentiation cultures, to study endoderm formation in the embryo, and to isolate pure populations of culture- or embryo-derived endodermal cells. Disclosure of potential conflicts of interest is found at the end of this article.Keywords
Funding Information
- NIH (U01DK072513)
- National Institute of Diabetes and Digestive and Kidney Diseases Grants (U01DK072477, U01DK072473)
This publication has 30 references indexed in Scilit:
- Anterior Definitive Endoderm from ESCs Reveals a Role for FGF SignalingCell Stem Cell, 2008
- Wnt, Activin, and BMP Signaling Regulate Distinct Stages in the Developmental Pathway from Embryonic Stem Cells to BloodCell Stem Cell, 2008
- A systematic screen for genes expressed in definitive endoderm by Serial Analysis of Gene Expression (SAGE)BMC Developmental Biology, 2007
- Sox17 Dependence Distinguishes the Transcriptional Regulation of Fetal from Adult Hematopoietic Stem CellsCell, 2007
- Wnt and TGF-β signaling are required for the induction of an in vitro model of primitive streak formation using embryonic stem cellsProceedings of the National Academy of Sciences of the United States of America, 2006
- Redundant roles ofSox17andSox18in postnatal angiogenesis in miceJournal of Cell Science, 2006
- The homeobox gene HEX regulates proliferation and differentiation of hemangioblasts and endothelial cells during ES cell differentiationBlood, 2005
- Embryonic stem cell differentiation: emergence of a new era in biology and medicineGenes & Development, 2005
- BMP Induction of Id Proteins Suppresses Differentiation and Sustains Embryonic Stem Cell Self-Renewal in Collaboration with STAT3Cell, 2003
- Continuous cultures of fused cells secreting antibody of predefined specificityNature, 1975