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N-2 Plus Media Supplement

Catalog #: AR003
24 Citations 2 Reviews Datasheet / COA / SDS
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AR003
Neural Progenitor Cells Expanded with N-2 Plus Media Supplement Express Nestin and SOX2
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Citations (24)
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N-2 Plus Media Supplement Summary

Kit Summary

For culturing neural progenitor cells and their differentiated derivatives.

Key Benefits

  • Optimized for NPC expansion and differentiation
  • Bovine Insulin is screened for batch-to-batch performance consistency
  • Chemically-defined and serum-free
  • Matures and maintains a variety of NPC-differentiated cell types
 

Why Culture Neural Progenitor Cells (NPCs) under Fully Defined Conditions?

Serum-free, defined media are routinely used as an alternative to standard serum-containing media in order to reduce unwanted experimental variability.

Uncontrolled variables commonly associated with serum-supplemented media, such as indeterminate levels of vitamins, hormones, and growth factors are eliminated, and the potential for contamination by infectious agents is reduced when cells are cultured under defined conditions. In addition, serum-free media offers researchers the ability to specifically design the culture media for a particular experimental question.

The N-2 Plus Media Supplement:

  • Contains high quality factors to support reproducible and efficient NPC expansion when combined with appropriate growth factors.
  • Is fully defined to reduce unwanted experimental variation.
  • Has been developed and optimized using neural cells.

N-2 Plus Media Supplement Components

Supplied as a 100X concentrate in water, this media supplement contains the following high quality factors to support neural cell culture:

  • Bovine Insulin (2,500 µg/mL)
  • Human Transferrin (10,000 µg/mL)
  • Putrescine (1,611 µg/mL)
  • Selenite (0.52 µg/mL)
  • Progesterone (0.63 µg/mL)

Supplied in a volume sufficient to supplement 500 mL of media at the recommended concentration.

 

Precautions

This product contains human transferrin. This transferrin was tested at the donor level using an FDA licensed method and found to be non-reactive for anti-HIV-1/2 and Hepatitis B surface antigen. As no testing can offer complete assurance of freedom from infectious agents, these reagents should be handled as if capable of transmitting infection.

Specifications

Shipping Conditions
The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at -20 to -70 °C. Use a manual defrost freezer and avoid repeated freeze-thaw cycles. Do not use past expiration date.

Product Datasheets

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Scientific Data

Immunocytochemistry Neural Progenitor Cells Expanded with N-2 Plus Media Supplement Express Nestin and SOX2 View Larger

Neural Progenitor Cells Expanded with N-2 Plus Media Supplement Express Nestin and SOX2 Neural Progenitor Cells Expanded with N-2 Plus Media Supplement Express Nestin and SOX2.Human neural progenitor cells were cultured for 7 days in media supplemented with 1X N-2 Plus Media Supplement (Catalog #AR003) and 20 ng/mL of Recombinant Human FGF basic (Catalog # 233-FB). The cells were stained with a PE-conjugated Mouse Anti-Human Nestin Monoclonal Antibody (Catalog# Catalog # IC1259P; red histogram), a PE-conjugated Mouse Anti-Human/Mouse SOX2 Monoclonal Antibody (Catalog# Catalog # IC2018P; green histogram), or a PE-conjugated Mouse IgG2AIsotype Control (Catalog# Catalog # IC003P; open histogram). Under these conditions, cells were shown to express high levels of Nestin and SOX2, two established markers of neural multipotency.

Immunocytochemistry Verification of Neural Progenitor Cell Multipotency Following Expansion with N-2 Plus Media Supplement View Larger

Verification of Neural Progenitor Cell Multipotency Following Expansion with N-2 Plus Media Supplement Verification of Neural Progenitor Cell Multipotency Following Expansion with N-2 Plus Media Supplement.Human neural progenitor cells were cultured for 7 days in media supplemented with 1X N-2 Plus Media Supplement (Catalog # AR003) and 20 ng/mL of Recombinant Human FGF basic (Catalog # 233-FB). Following the withdrawal of FGF basic, neural progenitor cells randomly differentiated into neurons, astrocytes, and oligodendrocytes. Markers of lineage differentiation were detected using a Mouse Anti-Human Nestin Monoclonal Antibody (Catalog # MAB1259), a Mouse Anti-Neuron-specific beta-III Tubulin (clone TuJ-1) Monoclonal Antibody (Catalog # MAB1195), a Sheep Anti-Human GFAP Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2594), and a Mouse Anti-Oligodendrocyte Marker O4 Monoclonal Antibody (Catalog # MAB1326). The cells were stained for Nestin using a NorthernLights trade; (NL)493-conjugated Donkey Anti-Mouse Secondary Antibody (Catalog # NL003; green), for beta-III Tubulin using a NL557-conjugated Donkey Anti-Mouse Secondary Antibody (Catalog # NL007; red), for GFAP using NL 557-conjugated Donkey Anti-Sheep Secondary Antibody (Catalog # NL010; red), and for O4 using an Anti-Mouse IgM secondary antibody (red). The nuclei were counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Stem Cells on Coverslips.

Assay Procedure

Refer to the product datasheet for complete product details.

Briefly, completed N-2 Plus-supplemented neural cell medium is prepared using the following procedure:

  • Dilute the media supplement in basal media
  • Store completed media
  • Use within 2 weeks
 

 

Reagents Provided

Reagents provided in the N-2 Plus Media Supplement (Catalog # AR003):

  • Bovine Insulin (2,500 µg/mL)
  • Human Transferrin (10,000 µg/mL)
  • Putrescine (1,611 µg/mL)
  • Selenite (0.52 µg/mL)
  • Progesterone (0.63 µg/mL)

 

Other Supplies Required

Reagents

  • DMEM/F-12 (Invitrogen®, Catalog # 12500-062) or a basal media (e.g., Neurobasal Media from Invitrogen, Catalog # 21103-029)
  • Glucose
  • Glutamine
  • NaHCO3
  • Penicillin-Streptomycin (100X)
  • Deionized or distilled water

Materials

  • Serological pipettes
  • Pipettes and pipette tips
  • 2 µm filter unit

Equipment

  • 2 °C to 8 °C refrigerator

 

Procedure Overview

Option 1: Mix the following components with deionized or distilled water to make 500 mL of medium.

Component Amount
DMEM/F-12 6 g
Glucose 0.775 g
Glutamine 0.0365 g
NaHCO3 0.854 g
N-2 Plus Media Supplement 5 mL

Adjust the pH to 7.2. Filter the solution (2 µm filter unit), and add 5 mL of 100X sterile Penicillin-Streptomycin solution. The medium may be stored in the dark at 2 °C to 8 °C for up to 2 weeks.

Option 2: Dilute 100-fold with a basal media (e.g., Neurobasal Media from Invitrogen, Catalog # 21103-029) before use. The medium may be stored in the dark at 2 °C to 8 °C for up to 2 weeks.

 

Invitrogen is a registered trademark of Invitrogen Corp.

Citations for N-2 Plus Media Supplement

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

24 Citations: Showing 1 - 10
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  1. Differential cell-cycle control by oscillatory versus sustained Hes1 expression via p21
    Authors: Maeda, Y;Isomura, A;Masaki, T;Kageyama, R;
    Cell reports  2023-05-17
  2. Lhx2 is a progenitor-intrinsic modulator of Sonic Hedgehog signaling during early retinal neurogenesis
    Authors: X Li, PJ Gordon, JA Gaynes, AW Fuller, R Ringuette, CP Santiago, V Wallace, S Blackshaw, P Li, EM Levine
    Elife, 2022-12-02;11(0):.  2022-12-02
  3. Improved modeling of human AD with an automated culturing platform for iPSC neurons, astrocytes and microglia
    Authors: R Bassil, K Shields, K Granger, I Zein, S Ng, B Chih
    Nature Communications, 2021-09-01;12(1):5220.  2021-09-01
  4. Ameliorative effects of aqueous extract of Forsythiae suspensa fruits on oxaliplatin-induced neurotoxicity in vitro and in vivo
    Authors: JM Yi, S Shin, NS Kim, OS Bang
    BMC Complement Altern Med, 2019-11-29;19(1):339.  2019-11-29
  5. Enhanced lysosomal degradation maintains the quiescent state of neural stem cells
    Authors: T Kobayashi, W Piao, T Takamura, H Kori, H Miyachi, S Kitano, Y Iwamoto, M Yamada, I Imayoshi, S Shioda, A Ballabio, R Kageyama
    Nat Commun, 2019-11-29;10(1):5446.  2019-11-29
  6. Mitochondrial transfer from mesenchymal stem cells to neural stem cells protects against the neurotoxic effects of cisplatin
    Authors: N Boukelmoun, GS Chiu, A Kavelaars, CJ Heijnen
    Acta Neuropathol Commun, 2018-12-12;6(1):139.  2018-12-12
  7. Reovirus-Induced Apoptosis in the Intestine Limits Establishment of Enteric Infection
    Authors: JJ Brown, SP Short, J Stencel-Ba, K Urbanek, AJ Pruijssers, N McAllister, M Ikizler, G Taylor, P Aravamudha, S Khomandiak, B Jabri, CS Williams, TS Dermody
    J. Virol., 2018-04-27;0(0):.  2018-04-27
  8. Zika virus infected primary microglia impairs NPCs proliferation and differentiation
    Authors: J Wang, J Liu, R Zhou, X Ding, Q Zhang, C Zhang, L Li
    Biochem. Biophys. Res. Commun., 2018-02-15;0(0):.  2018-02-15
  9. Activated CD8+ T lymphocytes inhibit neural stem/progenitor cell proliferation: role of interferon-gamma.
    Authors: Hu S, Rotschafer J, Lokensgard J, Cheeran M
    PLoS ONE, 2014-08-18;9(8):e105219.  2014-08-18
  10. Isolation of cancer stem cells from three human glioblastoma cell lines: characterization of two selected clones.
    Authors: Iacopino F, Angelucci C, Piacentini R, Biamonte F, Mangiola A, Maira G, Grassi C, Sica G
    PLoS ONE, 2014-08-14;9(8):e105166.  2014-08-14
  11. Proinflammatory cytokine-induced tight junction remodeling through dynamic self-assembly of claudins.
    Authors: Capaldo C, Farkas A, Hilgarth R, Krug S, Wolf M, Benedik J, Fromm M, Koval M, Parkos C, Nusrat A
    Mol Biol Cell, 2014-07-16;25(18):2710-9.  2014-07-16
  12. miR-17 regulates the proliferation and differentiation of the neural precursor cells during mouse corticogenesis.
    Authors: Mao S, Li H, Sun Q, Zen K, Zhang C, Li L
    FEBS J, 2014-01-15;281(4):1144-58.  2014-01-15
  13. Induction of intestinal stem cells by R-spondin 1 and Slit2 augments chemoradioprotection.
    Authors: Zhou, Wei-Jie, Geng, Zhen H, Spence, Jason R, Geng, Jian-Guo
    Nature, 2013-07-31;501(7465):107-11.  2013-07-31
  14. Transcription factor-mediated reprogramming of fibroblasts to expandable, myelinogenic oligodendrocyte progenitor cells.
    Authors: Najm, Fadi J, Lager, Angela M, Zaremba, Anita, Wyatt, Krysta, Caprariello, Andrew V, Factor, Daniel C, Karl, Robert T, Maeda, Tadao, Miller, Robert H, Tesar, Paul J
    Nat Biotechnol, 2013-04-14;31(5):426-33.  2013-04-14
  15. Matrix metalloproteinase-10 is required for lung cancer stem cell maintenance, tumor initiation and metastatic potential.
    Authors: Justilien V, Regala RP, Tseng IC
    PLoS ONE, 2012-04-24;7(4):e35040.  2012-04-24
  16. Directed differentiation of human pluripotent stem cells into intestinal tissue in vitro.
    Authors: Spence JR, Mayhew CN, Rankin SA
    Nature, 2010-12-12;470(7332):105-9.  2010-12-12
  17. Bmi1 is essential in Twist1-induced epithelial-mesenchymal transition.
    Authors: Yang MH, Hsu DS, Wang HW
    Nat. Cell Biol., 2010-09-05;12(10):982-92.  2010-09-05
  18. The giant protein AHNAK involved in morphogenesis and laminin substrate adhesion of myelinating Schwann cells.
    Authors: Salim C, Boxberg YV, Alterio J, Féréol S, Nothias F
    Glia, 2009-04-01;57(5):535-49.  2009-04-01
  19. Enhanced radiosensitivity and radiation-induced apoptosis in glioma CD133-positive cells by knockdown of SirT1 expression.
    Authors: Chang CJ, Hsu CC, Yung MC, Chen KY, Tzao C, Wu WF, Chou HY, Lee YY, Lu KH, Chiou SH, Ma HI
    Biochem. Biophys. Res. Commun., 2009-01-21;380(2):236-42.  2009-01-21
  20. Efficient Serum-Free Derivation of Oligodendrocyte Precursors from Neural Stem Cell-Enriched Cultures.
    Authors: Rao RC, Boyd J, Padmanabhan R, Chenoweth JG, McKay RD
    Stem Cells, 2009-01-01;0(0):.  2009-01-01
  21. Efficient induction of oligodendrocytes from human embryonic stem cells.
    Authors: Kang SM, Cho MS, Seo H, Yoon CJ, Oh SK, Choi YM, Kim DW
    Stem Cells, 2006-10-19;25(2):419-24.  2006-10-19
  22. Embryonic stem cell-derived neuron models of Parkinson's disease exhibit delayed neuronal death.
    Authors: Yamashita H, Nakamura T, Takahashi T, Nagano Y, Hiji M, Hirabayashi T, Amano T, Yagi T, Sakai N, Kohriyama T, Matsumoto M
    J. Neurochem., 2006-07-01;98(1):45-56.  2006-07-01
  23. Neuronal and glial expression of the adhesion molecule TAG-1 is regulated after peripheral nerve lesion or central neurodegeneration of adult nervous system.
    Authors: Soares S, Traka M, von Boxberg Y, Bouquet C, Karagogeos D, Nothias F
    Eur. J. Neurosci., 2005-03-01;21(5):1169-80.  2005-03-01
  24. High-level expression of functional chemokine receptor CXCR4 on human neural precursor cells.
    Authors: Ni HT, Hu S, Sheng WS, Olson JM, Cheeran MC, Chan AS, Lokensgard JR, Peterson PK
    Brain Res. Dev. Brain Res., 2004-09-17;152(2):159-69.  2004-09-17

FAQs

  1. What is the difference between N-2 Plus Media Supplement (Catalog # AR003) and N-2 MAX Media Supplement (Catalog # AR009)?

    • The N-2 Plus Media Supplement (Catalog # AR003) contains Bovine Insulin whereas N-2 MAX Media Supplement (Catalog # AR009) contains Recombinant Human Insulin. All other media components and concentrations in N-2 Plus and N-2 MAX are the same. These two products perform comparably in side-by-side testing. Due to a limited supply of bovine insulin, the products are priced differently.

  2. Can the N-2 MAX Media Supplement be thawed, aliquoted and refrozen to be used with smaller batches of basal media?

    •  Yes, this will not affect the stability of the supplement. Limited freeze-thaw of the supplement to use it in small batches is acceptable.

  3. Can the N-2 Plus Media Supplement be thawed, aliquoted and refrozen to be used with smaller batches of basal media?

    • Yes, this will not affect the stability of the supplement. Limited freeze-thaw of the supplement to use it in small batches is acceptable.

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N-2 Plus Media Supplement
By Leslie Priddy on 04/04/2018
N-2 Plus Media Supplement
By Anonymous on 12/01/2017

Great growth and stability of E14 Neuronal stem cells

N-2 Plus Media Supplement AR003