Human Induced Pluripotent Stem Cell (HiPSC) Line

Catalog Number: NGL-SC001
5 million and 10 million viable cells/vial

$1,250.00$2,500.00

Product Overview

NGL-SC001 HiPSC line derived from skin fibroblast of 18-year-old male donor. These cells are grown without antibiotics and maintain undifferentiated colonies for successful trilineage differentiation. NeyroblastGX HiPSC line is immune stable and has a high potential to generate any desired cells. We successfully generated neural progenitor cells, excitatory cortical neurons, microglial cells, cerebral organoids, choroid plexus organoids, intestinal organoids, and dendritic cells from the NGL-SC001 HiPSC line. The high-quality NGL-SC001 HiPSC line was reprogrammed using non-integrating reprogramming technology and was karyotypically stable and mycoplasma-free. One vial of HiPSC (>5 million cells/vial) contains large quantities of viable cells that can be directly used for downstream analysis and/or long-term culturing for multiple uses. NGL HiPSCs are optimized to grow in treated 6-well plates and 25.0 cm2 flasks in undifferentiated conditions. NGL-SC001 HiPSC line can be cultured on a robust scale, e.g.,  one vial of frozen viable HiPSC has the capability to culture a large number of 6-well plates or 25.0 cm2 flasks following our User Guideline.

NGL-SC001 HiPSC line derived from cells or tissues obtained via Institutional Review Board and/or an equivalent regulatory authority approved protocols. NGL-SC001 HiPSC line has undergone rigorous quality control and validation processes and demonstrates high levels of pluripotency marker expression to obtain any lineage cells.

Product Benefits

-High-quality undifferentiated NGL-SC001 HiPSC and capable of forming tightly packed colonies.

-Easy to maintain and passage HiPSC colony for long-term cultures.

-Ready to differentiate tri-lineage culture.

-Proven efficiency in differentiating ectodermal (neural progenitors, functional neurons, and cerebral organoids), mesodermal (dendritic and microglia cells), and endodermal (intestinal organoids) lineage cells. See related products for your study.

-Speed up your research with scalable quality control NGL-SC001 HiPSCs.

Storage, Stability, and Biosafety:

NGL-SC001 HiPSC lines are frozen in a cryopreservation medium CryoStor CS10, preformulated with 10% DMSO, providing a safe, protective environment during the freezing, storage, and thawing process for stem cells and tissues. NGL-SC001 HiPSC line is highly stable in liquid nitrogen (LN2) -130°C or colder and 12 months of date receipt. We strongly recommended storing the NGL-SC001 HiPSC line in LN2 to avoid undesirable differentiation in the culture. Thawed samples must be used immediately.

Caution: Please ensure the sample keeps storing LN2 (-130°C) upon receiving until further use. NeyroblastGX is not responsible if HiPSC lines do not work due to your LN2 temperature being compromised. The donor is free from viral or harmful pathogen infections. However, this product should treated as potentially hazardous materials and only used in appropriate biosafety level handling procedures such as those described in Biological Safety Level 2.

LIMITED USE LICENSE

NGL-SC001 HiPSC AND ITS DERIVED CELL PRODUCT ARE FOR RESEARCH USE ONLY AND ARE NOT INTENDED USE FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC PURPOSES OR COMMERCIALLY RESALED UNLESS OTHERWISE STATED. Please review the terms and conditions for the usage of NGL-SC001 HiPSC.

Characterization

NGL-SC001 HiPSC lines are maintained with cGMP grade medium without antibiotics; thus, HiPSCs passage provides high-quality, naïve, and undifferentiated colonies for obtaining desired lineages or cells shown in Figure 1.

 

Figure 1 update

Figure 1. NGL-SC001 HiPSC line was thawed and plated with ThermoFisher Scientific human recombinant Vitronectin coated plate and maintained with StemCell Technologies mTeSR plus media.  The white arrow showed a smooth edge of our undifferentiated HiPSC line. NGL-SC001 HiPSC line always show colonies with densely packed smooth edges (white arrow) with a high nucleus: cytoplasm ratio and viable bright cells (arrowhead). These morphological characteristics of HiPSC are crucial to obtaining successful differentiation and organoids, scale bar 150 µm.

L-SC001 HiPSC highly expresses undifferentiated pluripotent marker cells, as shown in Figure 2.

Figure 2 PRODUCT FIGURE 2

Figure 2. NGL-SC001 HiPSC line characterization: (A-C) High expression of SSEA4, OCT4, Tra-1-60, and SOX2 pluripotent marker cells by immunocytochemistry. (D) Flow cytometry determine Tra-1-81 expression in higher levels. (E) SOX2 and (F) NONOG mRNA expression of NGL-SC001 HiPSC line in compared to fibroblast (Fib) cells. Scale bar 100, 100 and 50 µm respectively.

L-SC001 HiPSC is a mycoplasma-free, karyotypically normal, and stable, as shown in Figure 3.

FIGURE 3

Figure 3. NGL-SC001 HiPSC maintained normal karyotype so that the HiPSC line will be used in any lineage-related differentiation cultures.

NGL-SC001 HiPSC characterized trilineage culture and successfully differentiated endodermal, mesodermal, and ectodermal cells, as shown in Figure 4.

Figure 4. NGL-SC001 HiPSC was differentiated into endodermal, mesodermal, and ectodermal lineage cells. (A-D) We have plated NGL-SC001 HiPSC with endodermal differentiation media, leading to a flat cell sheet with definite endodermal (DE) and hindgut spheroid precursor cells (arrowhead). DE showed robust expression of FOXA2 and SOX17 marker cells.  (E-H) We have also plated HiPSC for generating hematopoietic and myeloid cells. HiPSC-derived translucent myeloid was successfully differentiated into microglial and dendritic cells. Our HiPSC-derived early mesodermal cells express high-quality EMOES and CD45 marker cells. (I) NGL-SC001 HiPSC was coated with ThermoFisher Scientific Vitronectin and maintained with a neuroinduction medium to obtain neuroectoderm. We have observed a definite neuroectoderm with a neural tube-like rosette structure (arrowhead). (J) Neuroectoderm had further differentiated into neural progenitor cells with (K-L) high expression of SOX2, PAX6, and Nestin marker cells. Thus, NGL-SC001 HiPSC can differentiate into high-quality trilineage cells as needed for the project, scale bar 100-500 µm.

NGL-SC001 HiPSC successfully differentiated into neural progenitor cells (NPCs), as shown in Figure 5.

Figure 5. NGL-SC001 HiPSC-derived NGL-NPC005 NPCs neural progenitor cells characterization with the prominent NPC markers. (A) High expression of central nervous system (CNS) neural stem progenitor marker cells SOX2 (red) and Nestin (green) appears, which is determined by immunocytochemistry (ICC). (B) Robust expression of PAX6 (green) and (C) ZO1 (green) starts in neural stem/progenitor cells in the neural tube from initial CNS development and continues postnatal development as observed in NGL-SC001 HiPSC-derived NGL-NPC005 NPCs.  (D) Early expression of MAP2 (red) and the nestin (green) in higher amounts was also observed (blue = dapi nuclear stain, Scale bar 100 µm).

NGL-SC001 HiPSC successfully differentiated into excitatory cortical neurons to study corticogenesis and neurodegeneration, as shown in Figure 6.

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Figure 6. NGL-SC001 HiPSC-derived NGL-NPC005 NPCs differtiated into excitatory cortical neurons and their marker expression determined by immunocytochemistry. NPCs were cultured for 18 days with the outcome of a robust amount of (A) neuron-specific cytoskeletal protein markers MAP2 (red, scale bar 200 µm)(B) excitatory neuronal markers β III Tubulin and vesicular glutamate transporter 1 (VGLUT1, red, scale bar 25 µm), and (C-D) β III Tubulin (green) and deep layers of cortical neuronal marker T-Box Brain Transcription Factor 1 (TBR1, red) marker expressions (blue = dapi nuclear stain, Scale bar 25 µm).

NGL-SC001 HiPSC successfully differentiated into microglial cells to study glial-neuron interaction, as shown in Figure 7.

Figure 7. NGL-SC001 HiPSC line-derived healthy control microglia (NGL-MiCR008) highly expresses IBA1 to study neuron-glial interaction. (A) We first generated CD45 marker enrich microglial precursor cells (scale bar 100 µm) further treated with NeyroblastGX microglia complete growth media, which led to the outcome of (B) microglial type cells with vacuolated cytoplasm at day 7 (scale bar 100 µm). (C) On day 14, presented mature microglial morphology with many sinuous branches stemming from a small cell body (scale bar 70 µm), as shown in the insert image with a white arrowhead. (D) Microglial cells were immunostained with anti-IBA1 antibody and showed the robust outcome of IBA1 markers cells (scale bar 70 µm). (E) Flow cytometry of NGL-MiCR008 microglia histogram represents >70% IBA1 (green) marker cells and CD45 (blue) marker cells lower than 1% in the culture, which verified 99% of cells differentiated into microglia or microglia types cells using NGL microglia complete growth media.

NGL-SC001 HiPSC successfully differentiated into myeloid dendritic cells to study cell therapy, as shown in Figure 8.

Figure 8. Generation of myeloid DCs (NGL-DC007) from NGL-SC001 HiPSCs and validated its morphology and phagocytic activity. (A) Dissociated HiPSCs were treated with mesodermal growth factors to generate 3D spheroids and further generation of myeloid and immature dendritic cells (iDCs) with protrusion and veils type morphology. We treated iDCs with DC maturation factors to generate mature DCs (mDCs) with long dendrites. The arrowhead showed translucent myeloid, which time-dependently released iDC, but the arrow showed iDC with protrusion and veils type characteristics. Arrow also showed long processes in mDC, and Gimesa staining showed long dendrites. (B) We have tested DC’s phagocytic activity with FITC-IgG beads (green) for 24h, and DCs successfully engulfed beads >98%, which validates our DCs functionality using flow cytometry. Scale bars: 70-500 μm.

NGL-SC001 HiPSC successfully differentiated into Intestinal organoids to study microbiome versus pathobiome and intestinal disorders, as shown in Figure 9.

Figure 9. Formation of NGL-SC001 HiPSC-derived fully functional NGL-GIOr004 intestinal organoids. NGL spheroid, when plated with in-house gel matrix domes, enables the generation of a fully functional intestine by 18 days, (A) further maintaining a longer in culture 28 days fully mature with size ~3mm and the expression of (B) polarized inner endodermal and mesenchyme (CDX2+, green). (C) We also quantified crypt-type proliferating cells via Ki67 marker cells, which were determined with the comparison of nuclei (DAPI) in the same proliferating zone. We found that ~80% of our cells represent crypt-type cells. (D-F) On day 28, robust expression of endocrine cells (PDX1+, red) and highly proliferative SOX9+ (green), goblet cells (MUC2, red), and highly differentiated intestinal crypts proliferating cells (Ki67+, red). (G-I) ɑ-smooth muscle actin also expressed in our intestine, including enteroendocrine (ChromgA, chromogranin A+, green) and highly convoluted epithelial E-cadherin (E-cad+, red) and Paneth cell-derived lysozyme (red). All these expressions indicate our intestinal organoids are fully functional and capable of studying pathogens, microbiome, and chemically-induced intestinal toxicity, inflammation, bowel syndrome, or other intestinal disorders: scale bar 130 µm or higher, Nuclei, DAPI. The arrow indicates a specific marker expression.

Figure 10. NGL-SC001 HiPSC-derived mature NGL-COr003 cerebral organoids (OrGs) developed consisting of cortical layer neurons, glial cells and neuroepithelium. NGL cerebral OrG in the gel-matrix domes with the maintenance in a floating condition, leading to fully functional CNS cerebral OrG development within day 28. The NGL method to develop cerebral OrG required a time-dependent increase of complete neural growth media (NGL-hNGM005) in culture, capable of generating neuroepithelium, ventricular and subventricular zones (VZ and SVZ), and mature cortical layers neurons. For each stage of cerebral OrG formation, such as OrGspheroid to neural induction to cerebral OrGs, we have used the same neural media, unique to our system, for providing proper nutrition and integrating microglia without interrupting cortical patterning. Our cerebral OrG expresses (A-B) mature neuronal MAP2 (red) and astrocyte markers GFAP (green). Inserts showing visualize dendrites of neurons and spongiform morphology of astrocytes. (C) We have added cerebral OrG ready-to-use microglia, which efficiently integrated microglia with IBA1 (green) marker expression, (D) neuron-specific marker β III Tubulin (green), (E) late born superficial cortical layer marker (SATB2, green), (F) cortical deep layer marker FOXP2 (green), (G) cortical layers 2-3 marker BRN2 (green), and (H) cortical pre-plate to all the cortical layers marker TBR1 (red). (I) We have also observed epithelial tight junction protein ZO1 (green) marker expression. Our cerebral OrG is highly compatible with studying complex CNS diseases like Fragile X Syndrome, Autism Spectrum Disorders (ASD), Alzheimer’s, Parkinson’s’, and many other. Scale bar 130 µm, D; day.

User Guideline

HiPSC Culture Guideline

NGL-SC001 HiPSC line culture should be maintained with the procedure indicated in the User Guidelines. We will be sent a QR code for “User Guideline” upon ordering the product since each lot number of NGL-SC001 HiPSC frozen vial is unique. QR code is for downloading the digital copy of a detailed procedure for culturing and maintaining the NGL-SC001 HiPSC line. If you need a paper copy of the User Guidelines, please request it during ordering, and then we can ship it with NGL-SC001 HiPSC frozen vials.

Additional Information

Required Materials

NGL-SC001 HiPSC line culture maintenance and characterization will need the following associated materials but not included with our product: