Embryonic stem cells have the potential to differentiate into multiple cell
types and are widely recognized as holding significant promise for therapeutic
applications. The quality of a stem cell culture determines its ability to
give rise to differentiated cell types efficiently, and can be measured by
analysis of the expression of molecular markers of the pluripotent phenotype.
Markers of early lineage committed cells are also useful to determine whether
differentiated cells are present in a stem cell culture, and can indicate
which lineage pathway the differentiated cells represent.
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| Analytes |
- DNA marker
- Negative control
- DPPA5/ESG1
- Nanog
- Oct3/4
- SOX2
- Nestin
|
- Otx2
- TP63
- AFP
- GATA-4
- PDX-1
- SOX17
- HNF-3 beta
|
- Brachyury
- Stella
- GAPDH
- Positive control
- DNA marker
|
| Figure 1. PCR products amplified by primer sets specific
for 14 different markers of stem cell pluripotency and lineage commitment
visualized by gel electrophoresis. cDNAs used for amplification were
derived from a variety of embryonic tissues. Markers in lanes 3-6
are expressed in undifferentiated ES cells, lanes 6-9 ectodermal
lineage, lanes 10-14 endodermal lineage, lane 15 mesodermal lineage,
and lane 16 contains a marker for germ cells. Lanes 1, 2, and 17-19
show controls. |
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| Figure 2. Assessment of Oct3/4 expression in
NTera2 cells by intracellular flow cytometry using rat anti-human/mouse
Oct3/4 monoclonal antibody (Catalog # MAB1759;
orange) or isotype control (Catalog # MAB0061; gray) followed by
APC-conjugated secondary antibody. |
Figure 3. Detection of DPPA4 in the nuclei
of D3 cells using R&D Systems anti-mouse DPPA4 polyclonal antibody
(Catalog # AF3730) and
R&D Systems NorthernLights™ 557-conjugated secondary antibody
(Catalog # NL001). |
The R&D Systems Human Pluripotent Stem Cell Assessment Primer Pair Panel
(Catalog # SC012; Figure 1) profiles
the mRNA transcripts of fourteen genes that are frequently used as markers
for molecular characterization of undifferentiated and early lineage-committed
human ES cells. A primer pair for human GAPDH is included and can be used
as a control for successful cDNA synthesis. A positive control is also included.
In addition, R&D
Systems offers a mouse/rat version of this kit (Catalog # SC015).
Additional reagents to assess stem cell phenotype are available from R&D
Systems. For example, Oct 3/4 expression is detected in undifferentiated
NTera2 human teratocarcinoma cells by flow cytometry (Figure 2), and
DPPA4 expression is shown by immunocytochemistry in the nuclei of mouse D3
embryonic stem cells (Figure 3). |