Cells for Neural Stem Cell Research
ReNcell Immortalized Human Neural Progenitors and Serum-Free Media
- Convenient source of immortalized, human neural progenitor cells
- Immortalized cells replicate indefinitely, maintaining karyotype stability
- ReNcell Neural Progenitors are multipotent – they have been shown to differentiate into all three major neuronal lineages (neurons, astrocytes, oligodendrocytes)
- Readily expandable in proprietary, serum-free, defined ReNcell Media
- ReNcell VM Neural Stem Cell Line Models Parkinson’s Disease: New human disease model shows function of PINK1 gene in neural degeneration - Learn more...
Human Neural Stem Cells
ReNcell VM (SCC010) and CX (SCC009) are two established human neural stem cell lines, derived from the ventral mesencephalon and cortex brain regions of human fetal tissue* respectively, and immortalized by transduction with the transcription factor myc, to promote continuous growth and maintain a stable genotype and phenotype. ReNcells are the first easily available source of human fetal neural progenitors. They have been proven to grow for extended periods in culture (>45 passages), self-renew and differentiate into all three neuronal cell types.Multipotent Differentiation In Vitro
ReNcells are easily maintained in an undifferentiated state, showing the typical undifferentiated stem cell “cobblestone” morphology. Differentiation into multiple lineages is convenient and flexible, making these cells a valuable tool for a wide range of applications in basic stem cell biology, neuroscience and applied CNS-drug discovery fields. VM cells have been shown in vitro to differentiate into a high level of human dopaminergic neurons, shown to be electrophysiologically active. ReNcell CX can readily differentiate into neurons and glial cells.Stable, Normal Diploid Cells
The myconcogene appears to maintain normality and “stemness” of ReNcells in culture. In the presence of growth factors, ReNcells have been shown to maintain stable growth rates, karyotype, phenotype and multipotentiality, including specific neuronal phenotypes over prolonged passage. Upon removal of growth factors, cells rapidly differentiate into fully developed neurons.Proprietary, Serum-free, Defined Media
Optimized ReNcell Maintenance (SCM005) and Freezing Media (SCM007) provide a serum-free, defined environment for eliminating variables and ensuring the successful culture of ReNcells. *ReNcells have been isolated in a legal and ethical manner, compliant with local informed consent procedures.NEW ENStem-A™: Human Embryonic-derived Neural Progenitors and Optimized Media
ENStem-A Human Neural Progenitor Cells are derived from NIH approved WA09 human embryonic stem cells (hESCs). They proliferate as an adherent cell monolayer, maintain karyotype stability up to 10 passages and can readily differentiate into different neuronal subtypes. ENStem-A human neural progenitor cells may be used for a variety of research applications such as studies of neurotoxicity, neurogenesis, electrophysiology, neurotransmitter and receptor functions. ENStem-A cells are provided cryopreserved in a kit with 500 mls of Specially optimized Expansion Media containing FGF (SCR055). Also available separately are Expansion Medium (SCM004), Neuronal Differentiation Medium (SCM017) and Freezing Medium (SCM011).Mouse/Rat Neural Stem Cells
Neural Stem Cell lines include ready to use rat primary hippocampal neurons and astrocytes, primary neural stem cells isolated from the hippocampus of adult Fisher 344 rats, and mouse neural stem cells isolated from the cortices and spinal cord of embryonic day 15–18 (E15-E18) C57/BL6 mice. Each lot of primary cells has been validated for high expression of the appropriate markers, and with respect to adult rat NSC, for their self-renewal and multi-lineage differentiation capacity.Cutltured adult rat hippocampus-derived neural stem cells (SCR022) stained for (A) Nestin (red) and (B) Sox-2 (red). The Sox-2 transcription factor is colocalized with the DAPI (blue) staining in the nucleus. C. Mouse anti-Oligodendrocyte O1 (green) staining of adult rat hippocampus-derived neural stem cells (SCR022) that have been exposed to differentiation conditions for four days. (D) Localization of MAP-2 (orange) in primary rat hippocampus-derived embryonic neurons (SCR009, SCR010) that have been cultured for ten days. |
Neural Stem Cell Characterization Kits
To aid researchers in the accurate identification and characterization of NSCs and their differentiated progenies, Millipore offers the Neural Stem Cell Characterization Kit, the Neuron-Glial Cell Marker Sampler Kit and the ESC Derived Neuron Integration & Characterization Kit.The Neural Stem Cell Characterization Kit (SCR019) provides researchers with a convenient means to phenotype neural stem cells using a panel of antibodies. This kit contains reagents to the NSC markers Nestin and Sox-2, and also to markers of mature neural cells including neurons (Map2ab), astrocytes (GFAP) and oligoden- drocytes (Oligodendrocyte). Also included are mouse and rabbit immunoglobulins for the assessment of background staining.
The Neuron-Glial Marker Sampler Kit contains antibodies for the identification of neurons (ß-tubulin and Map2ab), astrocytes (GFAP) and oligodendrocytes (RIP). The ESC Derived Neuron Integration & Characterization Kit is a novel, broadly encompassing kit for the localization, characterization and analysis of ES cell-derived neuron integration into adult central nervous system (CNS) tissue. This kit allows researchers to classify mature neuronal pharmacological subtypes, and identify and quantify excitatory and inhibitory synapses involving the transplanted cells.
Adult Rat Hippocampal NSCs (SCR021, SCR022) are multipotent. NSCs were differentiated in either astrocyte (A-C) or neuronal and olygodendrocyte (D-F) differentiations medium. B. After nine days of differentiation in Astorcyte Differentiation Medium containing UF, BMP4, and fetal bovine serum (SCM010), the majority of the cells exhibit an astrocyte (GFAP, 85%) phenotype as assessed by positive immunoreactivity to astrocyte-specific marker, GFAP (red, C). E. NSCs cultured in the presence of retinoic acid and forskolin (SCR035) for five days differentiate predominately into Neurons (ß-tubulin, 70%; red, DJ and oligodendrocytes (O1, 30%; green, F). GFAP-positive astrocytes were not observed in this culture condition. Nuclei of the cells were visualized with DAPI (blue.) |
Rat Hippocampal Neural Stem Cell Kit
The Rat Hippocampal Neural Stem Cell Kit (SCR021) provides ready-to-use primary neural stem cells isolated from the hippocampus of adult Fisher 344 rats and antibodies for immunocytochemical staining of neural stem/progenitor cells (Nestin and Sox 2) and differentiated neural cells (MAP-2 for neurons, GFAP for astrocytes) for the culture and analysis of NSCs and their differentiated progenies. The viable, cryopreserved Rat Hippocampal Neural Stem Cells are also available separately.
Staining results obtained with rat hippocampal astrocyte kit components. Immunofluorescent image of primary astrocyte cells stained with rabbit andti-GFAP (red) and mouse anti-Map2ab (green). Neurons were not observed as evidenced by the lack of Map2ab staining. Nuclei of the cells were visualized with DAPI (blue, not included.) |
Rat Hippocampal Astrocyte & Neuron Kits
The Rat Hippocampal Astrocyte Kit and Rat Hippocampal Neuron Kit provide researchers with a convenient means for the culture and analysis of primary rat hippocampal astrocytes and neurons. In addition to viable, cryopreserved cells, these kits also include reagents for the immunostaining of astrocytes (Rabbit anti-GFAP antibody) and for the detection of neuron formation (Mouse anti-Map2ab antibody). Mouse and rabbit immunoglobulins for the assessment of background staining are also included. Alternatively, Cryopreserved Rat Hippocampal Astrocytes (SCR008) or Neurons (SCR010) are available as separate items.
The Neural Stem Cell Expansion Kits provide a multicomponent system for the culture and analysis of neural stem cells and their differentiated progenies. These systems include primary neural stem cells, neural stem cell expansion medium and a panel of antibodies for immunocytochemical staining of neural stem/progenitor cells (Nestin and Sox2) and differentiated neural phenotypes (Map2ab for neurons, GFAP for astrocytes and O1 for oligodendrocytes).
The Mouse Cortical Neural Stem Cell Expansion Kit (SCR032) includes mouse cortical neural stem cells, isolated from the cortices of embryonic day 15–18 (E15-E18) C57/BL6 mice; the Mouse Spinal Cord Neural Stem Cell Expansion Kit (SCR033) includes primary neural stem cells isolated from the spinal cord of embryonic day 15–18 (E15-E18) C57/BL6 mice; and the Adult Rat Neural Stem Cell Expansion Kit (SCR034) includes primary neural stem cells isolated from the hippocampus of adult Fisher 344 rats.
Rat Neural Stem Cell Blotting Kit
The Rat Neural Stem Cell Blotting Kit (SCR044) provides biotools that help monitor the expression of cell signaling molecules and proteins that are involved in the maintenance and differentiation of neural stem cells. Each kit contains two protein blots carrying normalized cellular extracts from neural stem cells and their differentiated progenies, neurons, astrocytes and oligodendrocytes. To assess the degree of protein normalization across cellular samples, an actin antibody is included. Also included are two neural-specific antibodies that are routinely used to monitor for the presence of neural stem cells (Nestin) and astrocytes (GFAP). As an extra added value, the ReBlot™ Plus Kit is also included to facilitate the multiple re-use of the protein blots.
Rodent Neuron Differentiation Kit
The Neuron Differentiation Kit (SCR035) provides two neuronal inducers that when added to the defined serum-free medium allow for the preferential differentiation of rodent neural stem cells to a neuronal lineage. The kit also includes antibodies for the immunocytochemical characterization of the resulting neuron population. It has been extensively validated on mouse cortical and spinal cord neural stem cells and on rat hippocampal neural stem cells. Using the Neuron Differentiation Kit, relatively pure populations of neurons (~70% ßIII-tubulin- positive) are obtained. Very low levels of astroglial cells (<0.5%) are detected.
Media for Neural Stem Cell Research
Adult rat hippocampal neural stem cells (SCR022) differentiated for 6-9 days in Astrocyte Differentiation Medium (SCM010). Over 90% GFAP-positive cells were detected with <1% Map2-positive neurons and <5% 01-positive oligodendrocytes. |
Free floating human neural stem cells (neurospheres) cultured in vitro using the growth factor hLIF in conjunction with bFGF and EGF (20x magnification) at initial passage (left) and 7-14 days later (right). The addition of hLIF improves long term expansion rates and results in an increased number of neurons produced following differentiation. Photographs courtesy of StemCells, Inc. |
Astrocyte Differentiation Medium
The Astrocyte Differentiation Medium (SCM010) is a specially formulated medium optimized for the preferential differentiation of rodent neural stem cells to an astrocyte lineage. The medium has been extensively validated on mouse cortical and spinal cord neural stem cells and on rat hippocampal neural stem cells. Using the Astrocyte Differentiation Medium, relatively pure populations of astrocytes (>80–90% GFAP-positive) are obtained. Very low levels of neurons (<1% MAP2AB-positive) or oligodendrocytes (<5% O1-positive) are detected.Neural Stem Cell Growth Factors
The conditions used to culture NSCs as free-floating neurospheres can impact on the number of neurons generated following differentiation. The number of neurons derived from human EGF-responsive neurospheres was shown to increase using specific culture conditions. This system utilized a cocktail of growth factors and permitted cell-to-cell contact during the differentiation stage (Carpenter, 1999, Experimental Neurology). These conditions resulted in an increased number of neurons produced (from 8% to >60%), with the most potent effects evident following the addition of Neurotrophic Factors 3 and 4 (NT3, NT4) and Platelet-Derived Growth Factor (PDGF). It was also demonstrated that the addition of Ciliary Neurotrophic Factor (CNTF) increased the number of astrocytes to 42%, compared to 20% derived from control cultures. Millipore’s comprehensive range of cytokines will facilitate the investigation of culture conditions for NSC expansion and factors influencing differentiation.Markers for Neural Stem Cells & Differentiated Progeny
Neural Stem Cell Markers
Millipore offers the largest line of neural markers available. Below, many interesting targets are called out.Undifferentiated Neural Stem Cells (NSCs) have been classically defined by the expression of the intermediate filament protein Nestin and are negative for the expression of markers of mature neural cells. In addition to these markers, numerous other antigens have been used to characterize NSCs.
Using a panel of antibodies to cell surface antigens, clonogenic NSCs were isolated from human fetal brain tissue by fluorescence-activated cell sorting (Uchida, 2000, PNAS). Purified NSCs were characterized as CD133+, CD34-, CD45-and CD24-/lo, and were able to engraft into the brains of immunodeficient neonatal mice. In another study, human neural progenitor cells isolated from postmortem human cortex were observed to express a number of markers previously associated with cultured fetal progenitor cells including CD24 (HSA), CD44 (Pgp-1), CD81, CD90 (Thy-1), CD133 and CD184 (Schwartz, 2003, J. Neuro. Res.).
Similarly, a number of antigens have been postulated as being useful for the identification of murine NSCs. A monoclonal antibody to Prominin-1 (CD133, human ortholog of the AC133 antigen) was used to show that this protein is expressed on the ventricular surface of the developing mouse mesencephalon, indicating that Prominin-1 could be used as a surface marker for neural precursor cells in mice (Sawamoto, 2001, J. Neurosci.). Using an alternative strategy, murine NSCs were isolated to 80% purity based on their unique FACS profile of markers that include Nestin, low level expression of CD24 (Heat Stable Antigen) and Peanut Agglutinin binding (Cai, 2004, J. Neurochemistry). Additionally, a population of adult murine NSCs was purified from the sub ventricular zone by FACS using a monoclonal antibody to the carbohydrate antigen SSEA-1 (Capela, 2002, Neuron).
Neural Stem Cell Markers
| BCRP1 | CD184 [C-X-X] CHEMOKINE RECEPTOR |
| CD24 [HEAT STABLE ANTIGEN] | NESTIN |
| CD34, CLASS I | NUCLEOSTEMIN |
| CD34, CLASS II | POLYSIALIC ACID-NCAM [PSA-NCAM] |
| CD34, CLASS iii | PROMININ-1 [CD133] |
| CD44S | SDNSF [NEURAL STEM CELL DERIVED NEURONAL SURVIVAL PROTEIN] |
| CD45 [LCA] | SOX-2 |
| CD81 [TAPA-1 | STAGE-SPECIFIC EMBRYONIC ANTIGEN-1 [SSEA-1] |
| CD90 [THY-1] |  |
Neural Stem Cell Differentiation Analysis
Once differentiated, NSCs have the potential to form the three major cell types of the central nervous system: neurons, astrocytes and oligodendrocytes. Antibodies specific for neuronal or glial proteins can is be utilized to identify the phenotypic properties of differentiated NSCs.Neuronal Markers
| AXONAL GROWTH CONES | NEUROFILAMENT 68-, 70-, 200-KD |
| CHOLINE ACETYLTRANSFERASE (ChAT) | NEURON SPECIFIC ENOLASE (NSE) |
| CHORMOGANIN A [CGA] | PERIPHERIN |
| DARPP-32 | PH8 |
| DOPAMINE TRANSPORTER [DAT] | PROTEIN GENE PRODUCT 9.5 |
| DOPA DECARBOXYLASE | SEROTONIN TRANSPORTER (SERT) |
| GABA | SYNAPSIN 1 |
| GLUTAMATE DECARBOXYLASE | SYNAPTOPHYSIN |
| GROWTH ASSOCIATED PROTEIN 43 | TAU |
| HUC | TRK A |
| HUD | TRYPTOPHAN HYDROXYLASE |
| INTERNEXIN A | ß11 TUBULIN |
| MAP1B [MAP5] | TYROSINE HYDROXYLASE |
| MAP2A, 2B | VANILLOID RECEPTOR-LIKE PROTEIN [VRL-1] |
| MAP2 | VESICULAR GABA TRANSPORTER |
Astrocyte Markers
| GFAP |
| S-100 protein |
Oligodendrocyte Markers
| A2B5 | CNPASE |
| GALACTOCEREGROSIDE | MYELIN BASIC PROTEIN |
| NG2 CHONDROTIN SULFATE PROTEOGLYCAN | 01 |
| 04 | OLIGODENDROCYTES |
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