N21-MAX Media Supplement (50X) Summary
For the reliable maturation and superior long-term culture of primary neurons.
- Provides reliable maturation and superior function of neurons in culture
- Can be used to improve stem cell and 3D culture conditions
- Rigorous quality control testing ensures consistency across experiments
- Is nutrient-rich and serum-free
- Eliminates the need for an astrocyte feeder layer in neuronal cultures
Why Culture Neurons in Serum-Free N21-MAX Media Supplement?
The N21-MAX Media Supplement improves upon the traditional B271 and NS212 neuronal supplements, offering a serum-free and fully-defined formulation that is optimized for the reliable maturation, consistent health, and superior function of neurons in culture. There is an increase in the use of B27-like supplements to optimize stem cell and 3D cell culture media. N21-MAX improves stem cell differentiation and increases the viability and health of differentiated cell types during long-term cell culture conditions.
N21-MAX was designed to eliminate uncontrolled variables, such as those found in both serum-containing media and in some commercial serum-free neuronal supplements. Use of undefined media supplements can confound neuronal growth and maturation. N21-MAX contains a cocktail of factors optimized for neuronal survival, neurite extension, and synaptic maturation. Each lot of N21-MAX is tested on primary hippocampal neurons to ensure consistent performance for the user.
The N21-MAX Media Supplement:
- Is fully defined and quality tested to reduce unwanted experimental variation.
- Contains factors qualified to support reproducible and long-term neuron culture.
- Modified from the published B27 formulation1.
- Does not require an astrocyte feeder layer.
- Has been tested to support the viability and growth of E18 rat hippocampal neurons.
- Brewer et al., (1993) J. Neurosci. Res. 32:567
- Chen et al., (2008) J. Neurosci. Methods 171:239
N21-MAX is optimized for the maturation and long-term culturing of neuronal cells. The supplement is supplied as a 50X concentrated solution and contains the following 21 components:
- Albumin (bovine)
- Linoleic Acid
- Linolenic Acid
- Lipoic Acid
- Retinyl acetate
- Superoxide dismutase
- D,L-alpha-Tocopherol acetate
Supplied in a volume sufficient to supplement 500 mL of media at the recommended concentration. N21-MAX is tested for use in Neurobasal Media.
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.
N21-MAX Reduces Spontaneous Stress Rod Formation. E18 rat hippocampal neuron cultures were plated on poly-D-lysine coated coverslips and grown for 5 days in media supplement with either N21-MAX (Catalog # AR008) or another commercial neural media supplement (Competitor). Cells were fixed and immunostained for rod formation, and indicator of cellular stress, using an antibody against cofilin (% of Neurons with Rods). A lower percentage of hippocampal neurons that were cultured in N21-MAX had spontaneous cofilin rod formation (4.5%) compared to neurons cultured in competitor media (16.8%). Data courtesy of the laboratory of Dr. James Bamburg at Colorado State University.
Improved Resolution of Rod Induction in Neurons Cultured in N21-MAX. E18 rat hippocampal neuron cultures were plated on poly-D-lysine coated coverslips and grown for 5 days in N21-MAX (Catalog # AR008). Neurons were then treated with rod-inducing agents or maintained in N21-MAX alone (Untreated). Neurons were fixed and immunostained for rod formation using an antibody against cofilin (% of Neurons with Rods). The low background of rod formation in untreated cultures enabled detection of mild rod induction using Synthetic Amyloid beta, Amyloid beta dimer/trimer, and TNF-alpha. As a positive control, treatment with an ATP depletion solution or glutamate resulted in robust rod induction. Data courtesy of the laboratory of Dr. James Bamburg at Colorado State University.
N21-MAX Media Supplement Enhances Synaptic Development. E18 rat hippocampal neurons were grown for 19 days in vitro in media supplemented with either N21-MAX Media Supplement or the neural media supplement from the most widely-used competitor. Cells were incubated with the synaptic vesicle dye, SynaptoRed™C2 (10 µM; Catalog # 5118), for 1 minute prior to depolarization with 50 mM KCl. Cells were imaged for SynaptoRed™C2-positive synaptic puncta using the Operetta CLS™High Content Analysis System. (A) Quantification of SynaptoRed™C2-positive puncta shows that neurons grown in N21-MAX have an increased number of synaptic puncta compared to the competitor media. (B) Quantification of dye intensity shows that neurons grown in N21-MAX have more robust synaptic activity than neurons cultured in the competitor media. (C) Representative images of SynaptoRed™-C2 staining in neurons cultured in N21-MAX or competitor media. (D) Representative images showing the quantification of synaptic puncta in neurons cultured in N21-MAX or competitor media. Colored circles indicate a SynaptoRed™-positive synaptic puncta.
Synaptotagmin and CAM Kinase II Expression in Neurons Cultured in Media Supplemented with N21-MAX. Synaptotagmin and CAM Kinase II Expression in Neurons Cultured in Media Supplemented with N21-MAX.E18 rat hippocampal neurons were grown for 21 daysin vitroin media supplemented with N21-MAX Media Supplement (Catalog # AR008). Cells were stained with either (A) a Mouse Anti-Rat Synaptotagmin-1 Monoclonal Antibody (Catalog # MAB4364), followed by the NorthernLights™(NL)557-conjugated Donkey Anti-Mouse IgG Secondary Antibody (Catalog # NL007; yellow), or (B) a Mouse Anti-Human CaM Kinase II alpha Monoclonal Antibody (Catalog # MAB5584), followed by the NL557-conjugated Donkey Anti-Mouse IgG Secondary Antibody (Catalog # NL007; green). The nuclei were counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Increased Synaptic Puncta and Neurite Outgrowth of Primary Neurons Cultured in N21-MAX. Increased Synaptic Puncta and Neurite Outgrowth of Primary Neurons Cultured in N21-MAX.E18 rat hippocampal neurons were grown for 21 daysin vitroin media supplemented with either N21-MAX Media Supplement (Catalog # AR008) or the neural media supplement from the most widely-used competitor. Staining for Synaptotagmin (yellow) showed more robust synaptic puncta and increased neurite outgrowth in neurons cultured in N21-MAX compared to those cultured in competitor media. Cells were staineed with a Mouse Anti-Rat Synaptotagmin-1 Monoclonal Antibody (Catalog # MAB4364) followed by the NorthernLights™(NL)557-conjugated Donkey Anti-Mouse IgG Secondary Antibody (Catalog # NL007). Nuclei were counterstained with DAPI (blue).
N21-MAX Media Supplement Increases Efficiency of Pancreatic Cell Differentiation. N21-MAX Media Supplement Increases Efficiency of Pancreatic Cell Differentiation.Differentiation of PDX-1+pancreatic cells from induced pluripotent stem cells (iPS2) was performed with base media containing either the N21-MAX Media Supplement (green) or the competitor equivalent (red).A) Compared to competitor media, N21-MAX improved pancreatic cell differentiation as observed by flow cytometry.B) Bar graph quantifying PDX-1+cells in N21-MAX and the competitor media.
Refer to the product datasheet for complete product details.
Briefly, completed N21-MAX-supplemented medium is prepared using the following procedure:
- Dilute the media supplement in basal media
- Store completed media
- Use within 2 weeks
Reagents provided in the N21-MAX Media Supplement (Catalog # AR008):
- Albumin (bovine)
- Linoleic Acid
- Linolenic Acid
- Lipoic Acid
- Retinyl acetate
- Superoxide dismutase
- D,L-alpha-Tocopherol acetate
Other Supplies Required
- Basal media (e.g., Neurobasal Media from Invitrogen®, Catalog # 21103-029 or equivalent)
- Serological pipettes
- Pipettes and pipette tips
- 2 °C to 8 °C refrigerator
Dilute 50-fold with a basal media (e.g., Neurobasal Media from Invitrogen, Catalog # 21103-029) and supplement with 0.5 mM L-glutamine 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 N21-MAX Media Supplement (50X)
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.
Citations: Showing 1 - 10
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Cardiac glycoside-mediated turnover of Na, K-ATPases as a rational approach to reducing cell surface levels of the cellular prion protein
Authors: M Mehrabian, X Wang, S Eid, BQ Yan, M Grinberg, M Siegner, C Sackmann, M Sulman, W Zhao, D Williams, G Schmitt-Ul
PLoS ONE, 2022;17(7):e0270915. 2022
Cis- and trans-resveratrol have opposite effects on histone serine-ADP-ribosylation and tyrosine induced neurodegeneration
Authors: M Jhanji, CN Rao, JC Massey, MC Hope, X Zhou, CD Keene, T Ma, MD Wyatt, JA Stewart, M Sajish
Nature Communications, 2022;13(1):3244. 2022
Impact of schizophrenia GWAS loci converge onto distinct pathways in cortical interneurons vs glutamatergic neurons during development
Authors: D Liu, A Zinski, A Mishra, H Noh, GH Park, Y Qin, O Olorife, JM Park, CP Abani, JS Park, J Fung, F Sawaqed, JT Coyle, E Stahl, J Bendl, JF Fullard, P Roussos, X Zhang, PK Stanton, C Yin, W Huang, HY Kim, H Won, JH Cho, S Chung
Molecular Psychiatry, 2022;0(0):. 2022
Lysosomal exocytosis releases pathogenic alpha-synuclein species from neurons in synucleinopathy models
Authors: YX Xie, NN Naseri, J Fels, P Kharel, Y Na, D Lane, J Burré, M Sharma
Nature Communications, 2022;13(1):4918. 2022
CNS myelination requires VAMP2/3-mediated membrane expansion in oligodendrocytes
Authors: M Lam, K Takeo, RG Almeida, MH Cooper, K Wu, M Iyer, H Kantarci, JB Zuchero
Nature Communications, 2022;13(1):5583. 2022
Regulation of host and virus genes by neuronal miR-138 favours herpes simplex virus 1 latency
Authors: B Sun, X Yang, F Hou, X Yu, Q Wang, HS Oh, P Raja, JM Pesola, EAH Vanni, S McCarron, J Morris-Lov, AHM Ng, GM Church, DM Knipe, DM Coen, D Pan
Nature Microbiology, 2021;0(0):. 2021
The cellular prion protein interacts with and promotes the activity of Na,K-ATPases
Authors: D Williams, M Mehrabian, H Arshad, S Eid, C Sackmann, W Zhao, X Wang, F Ghodrati, CE Verkuyl, JC Watts, G Schmitt-Ul
PLoS ONE, 2021;16(11):e0258682. 2021
Direct interaction of HIV gp120 with neuronal CXCR4 and CCR5 receptors induces cofilin-actin rod pathology via a cellular prion protein- and NOX-dependent mechanism
Authors: LK Smith, IW Babcock, LS Minamide, AE Shaw, JR Bamburg, TB Kuhn
PLoS ONE, 2021;16(3):e0248309. 2021
Silencing of AURKA augments the antitumor efficacy of the AURKA inhibitor MLN8237 on neuroblastoma cells
Authors: Y Yang, L Ding, Q Zhou, L Fen, Y Cao, J Sun, X Zhou, A Liu
Cancer Cell Int., 2020;20(0):9. 2020
Live Neuron High-Content Screening Reveals Synaptotoxic Activity in Alzheimer Mouse Model Homogenates
Authors: H Jiang, TJ Esparza, TT Kummer, H Zhong, J Rettig, DL Brody
Sci Rep, 2020;10(1):3412. 2020
Neural stem cell small extracellular vesicle-based delivery of 14-3-3t reduces apoptosis and neuroinflammation following traumatic spinal cord injury by enhancing autophagy by targeting Beclin-1
Authors: Y Rong, W Liu, C Lv, J Wang, Y Luo, D Jiang, L Li, Z Zhou, W Zhou, Q Li, G Yin, L Yu, J Fan, W Cai
Aging (Albany NY), 2019;11(18):7723-7745. 2019
Tau interactome analyses in CRISPR-Cas9 engineered neuronal cells reveal ATPase-dependent binding of wild-type but not P301L Tau to non-muscle myosins
Authors: X Wang, D Williams, I Müller, M Lemieux, R Dukart, IBL Maia, H Wang, AL Woerman, G Schmitt-Ul
Sci Rep, 2019;9(1):16238. 2019
Stemness-Attenuating miR-503-3p as a Paracrine Factor to Regulate Growth of Cancer Stem Cells
Authors: M Seo, SM Kim, EY Woo, KC Han, EJ Park, S Ko, EW Choi, M Jang
Stem Cells Int, 2018;2018(0):4851949. 2018
Axonal G3BP1 stress granule protein limits axonal mRNA translation and nerve regeneration
Authors: PK Sahoo, SJ Lee, PB Jaiswal, S Alber, AN Kar, S Miller-Ran, EE Taylor, T Smith, B Singh, TS Ho, A Urisman, S Chand, EA Pena, AL Burlingame, CJ Woolf, M Fainzilber, AW English, JL Twiss
Nat Commun, 2018;9(1):3358. 2018
Development of a central nervous system axonal myelination assay for high throughput screening.
Authors: Lariosa-Willingham K, Rosler E, Tung J, Dugas J, Collins T, Leonoudakis D
BMC Neurosci, 2016;17(0):16. 2016
Silencing of AURKA augments the antitumor efficacy of the AURKA inhibitor MLN8237 on neuroblastoma cells.
Authors: Yang Y, Ding L, Zhou Q, Fen L, Cao Y, Sun J, Zhou X, Liu A
Cancer Cell Int, 0;20(0):9. 0
What advantages does the N21-MAX Media Supplement (Catalog # AR008) have over other neuronal stem cell supplements?
We have performed comparison testing in-house and have found that in long term cultures (21 days), we observe better synapse marker expression.
Additionally, one major formulation difference is that our N21 contains holo-transferrin while other supplements typically contain apo-transferrin. It is reported in the literature that holo-transferrin is preferred for neuronal cell culture. We have also tested several BSA sources to determine which is optimal for neuronal cultures.
Reviews for N21-MAX Media Supplement (50X)
Average Rating: 4.3 (Based on 6 Reviews)
Have you used N21-MAX Media Supplement (50X)?
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The area, intensity, and density of PSD-95 signal are comparable to the culture grown in B27.
Adult human stem cells showed fast growth and took less time to differentiate into neurons. Data is communicated so cannot upload image.
I have previously used Gibco B27®, and Gem21 NeuroPlex™ for culturing of neural cells and neuronal lines. I use typical for co- culture of neurons with astrocytes and microglia . I tried your sample N21 max from R&D and it worked very well with my neuronal cells co cultures. All of these Gibco B27®, and Gem21 NeuroPlex™ and N21 max are Supplied as a 50X concentrate and are list priced at $79 for 10 ml. I would use them all interchangeably as they all worked equally well in my co culture system.
For stem cell culture media supplementation.