iPS Cells (iPSC from Fibroblasts, Episomal, Male, Control Lines for Genome Editing Services); NIST Line *Academic Price - Research

SKU :
ASE-9211
Catalog # :
ASE-9211
Size :
0.5 x 10^6 cells/vial
$2,500.00
Description

MTA required

Applied StemCell, Inc. provides Control Human Induced Pluripotent Stem Cells (iPSC) at low passages (p15). These pluripotent cells were generated from normal human skin fibroblasts using episomal reprogramming methods.  This method allows the transient expression of human transcription factors (OCT4, SOX2, KLF4, and c-MYC) that initiate the reprogramming process. The resulting human iPS cells (hiPSCs) were selected using morphological criteria without the use of either fluorescent markers or drug selection. These iPS cells have been tested for the expression of the pluripotency markers, including OCT4, SOX2, SSEA4, TRA-1-60, TRA-1-81, and alkaline phosphatase (AP) activity, and normal male karyotype. The ASE-9211 control human iPSC line has proven capability for CRISPR/Cas9 genome editing and differentiation to various somatic lineages.

Detailed protocols for thawing, culturing under feeder-free conditions, and cryopreservation of these iPS cells are provided.

Applied StemCell (ASC) is a member of the NIST Genome Editing Consortium and our  ASE-9211 “Master” iPSC line is being used as a benchmark material for developing measurements and standards for the rapidly evolving genetically engineered cells used in cell therapy research.

For donor information, please refer to datasheet or contact us.


Characterization

Pluripotency Marker Analysis

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Figure 1. Expression of pluripotency markers. ASC 9211 iPS cell line expresses common iPSC biomarkers (top row: OCT-4, SOX2, SSEA-4, TRA-1-60, and TRA-1-81).  Bottom row: Corresponding DAPI nuclear staining.  All images were taken at 10x magnification.

Alkaline Phosphatase (AP) Staining

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Figure 2. Alkaline Phosphatase (AP) staining. ASC-9211 iPSCs stain positive for Alkaline Phosphatase: a typical unstained colony (a) was used to gauge the extent of the AP staining (b). Both images were taken at 5x magnification.

Karyotype Analysis

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Figure 3. Karyotype analysis to rule out genetic aberrations. This iPSC line demonstrates a normal male karyotype. Cytogenic analysis was performed on twenty G-banded metaphase cells from human iPSC line, ASE-9211 at passage 15. Nineteen cells demonstrated an apparently normal male karyotype, and one cell demonstrated a non-clonal chromosome aberration, which is most likely an artifact of culture.

Karyotype Analysis

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Figure 4. Quantitative PCR (qPCR) analysis to detect karyotypic abnormalities in control human iPSC line, ASE-9211 shows a normal male karyotype in all loci tested.

Directed-differentiation to Three Germ Layers

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Figure 5. Direct differentiation of ASE-9211 p18 to three germ layers. Immunofluorescent staining for lineage-specific biomarkers of three germ layers after direct differentiation of control hiPSC line, ASE-9211. The ASE-9211 hiPSC line was differentiated to specific lineages of the germ layers using well-established and optimized protocols. Immunostaining for biomarkers of each lineage was performed to confirm lineage commitment. Cells were also co-stained with nuclear marker, DAPI (blue). Images shows are co-localization of biomarker with DAPI. Endoderm (EN) marker: SOX17 (green); Mesoderm (ME) marker: GATA4 (green); Ectoderm (EC) marker: b-III Tubulin (red).


CRISPR Genome Engineering Capability

 

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Figure 6. CRISPR/Cas9 was used to generate 6 knockout cell lines from the control human iPSC line, ASE-9211.


Differentiation Capability

Control iPSC line, ASE-9211 was used for differentiation into motor neurons using proprietary, integration-free protocols. 

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Figure 7. Immunocytochemical staining images of the motor neuron derived from control "master" iPSC line, ASE-9211 at 2 days post thaw. The iPSC-derived motor neurons were stained with antibodies for motor neuron biomarker, HB9 (green), and neuronal biomarker, Tuj1 (red). DAPI (blue) was used as nucleus stain.

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Figure 8. Immunocytochemical staining for motor neuron marker, ChAT (green) and neuronal marker, MAP2 i(blue) n motor neurons differentiated from control "master" iPSC line, ASE-9211 at 7 days post thaw. DAPI (blue) was used for nucleus staining.


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