B-27 Plus Neuronal Culture System

The standard in neuronal cell culture just got better

Cell survival rates are critical to the success and reliability of your neurobiology research. The Gibco B-27 Plus Neuronal Culture System, comprised of B-27 Plus Supplement and Neurobasal Plus Medium helps improve the reliability of your experiments by increasing the survival of neurons by more than 50%.

See what researchers are saying 

  • Increased neuronal survival by more than 50%—the highest in the industry
  • Accelerated neurite outgrowth
  • Improved electrophysiological activity and maturation of neurons
     

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Gibco B-27 Plus Neuronal Culture System includes

Gibco B-27 Plus Neuronal Culture System includes B-27 Plus Supplement and Neurobasal Plus Medium.

Validated to work best together—Order the convenient and valuable all-in-one system and re-order individual components as needed

Proven to seamlessly replace other neuronal cell culture systems, such as classic regular versions of B-27 Supplement and Neurobasal Medium, with no changes to current workflows for:

  • Maintenance of primary rat, mouse and human PSC-derived and fetal-derived neurons
  • Differentiation of human PSC-derived and fetal-derived Neural Stem Cells (NSC) to neurons
     

Key benefits and representative data

Designed to increase neuronal survival by more than 50% compared to the classic products

The B-27 Plus Culture System increases neuronal survival over other neuronal cell culture systems.

Figure 1. B-27 Plus Neuronal Culture System maintains higher neuronal survival rates compared to B-27 Supplement and Neurobasal Medium. Gibco Primary Rat Cortex Neurons were cultured for up to 3 weeks in the listed media system and then immunostained for MAP2 on days 7, 14, and 21.

B-27 Plus Neuronal Culture System enables superior cultures of primary and iPSC-derived neurons.

Figure 2. B-27 Plus Neuronal Culture System enables superior cultures of primary rodent and induced pluripotent stem cell (iPSC)–derived neurons compared to B-27 Supplement and Neurobasal Medium. Cryopreserved neurons were cultured for 3–4 weeks in the listed media system. Neurons were immunostained at the indicated time points for the neuronal dendritic marker MAP2 (green) and the neuronal cell body marker HuC/D (red). Nuclei were counterstained with DAPI (blue).

Improves electrophysiological activity and maturity

B-27 Plus Neuronal Culture System is exhibits highly synchronized neuronal activity over time.

Figure 3. Improvement of electrophysiological activity compared to BrainPhys Neuronal Medium and SM1 Kit. Primary Rat Cortex Neurons plated on 48-well multi-electrode array (MEA) plates. Cells were cultured for 35 days in the listed media system following the suppliers’ recommended protocols. Spontaneous electrophysiological activity was recorded throughout with the Axion BioSystems™ Maestro™ MEA platform. Data shown are from one of four experiments, with each run showing similar trends. The B-27 Plus Neuronal Culture System enabled more consistent, stable, and highly synchronized spontaneous electrophysiological activity over time.

Enhanced neuronal maturation within healthy neuronal cell culture.

Figure 4. Enhancement of neuronal maturation compared to B-27 Supplement and Neurobasal Medium. Primary Rat Cortex Neurons at day 22 were stained for the dendritic marker MAP2 (red) and synapsin 1/2 to label presynaptic terminals (green). Neurons maintained in the B-27 Plus Neuronal Culture System had significantly higher synaptic-positive puncta.

B-27 Plus Neuronal Culture System promotes increased neural network activity.

Figure 5. The B-27 Plus Neuronal Culture System promotes increased neural activity. Primary rat cortex neurons were maintained for 35 days in vitro in B27 Plus system with half-medium changes every 2–3 days. Recordings were acquired 24 hours after media changes using Axion BioSystems Maestro Pro™ microelectrode array (MEA) platform. The B-27 Plus system exhibited neural coverage across the entire array (see activity map, left) and strong, synchronized network phenotype, as illustrated in the raster plot (activity map, right). (Data generated by Axion Biosystems.

Supports highest survival of neurons compared to alternative media systems

Figure 6. The B-27 Plus Neuronal Culture System achieves the highest neuronal survival in long-term cultures compared to other commercially available serum-free neuronal media systems. Primary Rat Cortical Neurons, Primary Rat Hippocampus Neurons, Primary Mouse Cortical Neurons and human iPSC-derived neurons that had been matured for 9 days before cryopreservation (HIP™ Neurons, MTI-GlobalStem) were thawed in classic Neurobasal Medium with B-27 Supplement. The neurons were plated onto poly-D-lysine–coated 96-well plates and maintained for 3–4 weeks in the B-27 Plus Neuronal Culture System and alternative serum-free supplemented media systems following the suppliers’ recommended protocols. Neuronal survival was quantitated by immunofluorescent labeling using Invitrogen HuC/HuD Monoclonal Antibody at 3 or 4 weeks. Data shown are from one of three experiments, with each run showing that B-27 Plus Neuronal Culture System enables the highest neuronal cell survival among the alternative systems.

Figure 7. Acceleration of neurite outgrowth compared to B-27 Supplement and Neurobasal Medium and alternative media systems. Primary Mouse Cortical Neurons were thawed in classic Neurobasal Medium with B-27 Supplement and plated onto poly-D-lysine–coated 96-well plates. Neurons were maintained for ~3 weeks in Neurobasal Medium with B-27 Supplement, Neurobasal Plus Medium with B-27 Plus Supplement, and other commercially available serum-free neuronal media systems following the suppliers’ recommended protocols. Neurite outgrowth was quantitated using differential interference contrast images taken at the time points specified. The B-27 Plus Neuronal Culture System significantly accelerates neurite outgrowth over the first few weeks compared to classic Neurobasal Medium with B-27 Supplement. Data shown are from one of three experiments, with each run showing after 1 week B-27 Plus Neuronal Culture System enabled the highest mean neurite length compared to all alternative systems.

Customer stories

Primary mouse hippocampal neruons (MHN) were cultivated over a period of 28 days with Neurobasal Plus Medium supplemented with B-27 Plus Supplement under standard conditions. The cells showed healthy morphology and no neurite fragmentation could be observed. All dendritic spine morphologies that can be found in vivo, could also be observed in cell culture. I would clearly recommend the Neurobasal Plus Medium and B-27 Plus Supplement for the long time cultivation of primary neurons.

— Susanne Zach, Boehringer Ingelheim, CNS Research

Susanne Zach
Professor Hideyuki Okano
The neurons in the B-27 Plus system appeared healthier, with longer and more complete neurites, as compared to the other media conditions. We are extremely happy with this new media system and plan on continuing to use it in our experiments as we believe that it will allow us to more efficiently and effectively study the differentiation and maturation of NSC and iPSC-derived neurons.

—  Professor Hideyuki Okano, MD, PhD, Keio University School of Medicine, Tokyo, Japan

With our primary rat cortical neural cultures, the B-27 Plus Neuronal Culture System produces robust neural activity across almost every electrode in MEA plates. The network bursting events are very regular, and extremely high intensity, making data analysis easy and straightforward when evaluating functional neural activity.

— Jim Ross, CTO at Axion BioSystems

Jim Ross
John Graef
B-27 Plus medium led to superior levels of spontaneous multielectrode array activity in our human iPSC-derived neurons as compared to our standard preparations with BrainPhys culture media. I found that B-27 Plus medium represents an excellent option for cell culture systems where identifying potential electrophysiological phenotypes based on spontaneous neuronal activity is desired.

— John Graef, PhD, Principal Scientist, Fulcrum Therapeutics

Designed to increase neuronal survival by more than 50% compared to the classic products

The B-27 Plus Culture System increases neuronal survival over other neuronal cell culture systems.

Figure 1. B-27 Plus Neuronal Culture System maintains higher neuronal survival rates compared to B-27 Supplement and Neurobasal Medium. Gibco Primary Rat Cortex Neurons were cultured for up to 3 weeks in the listed media system and then immunostained for MAP2 on days 7, 14, and 21.

B-27 Plus Neuronal Culture System enables superior cultures of primary and iPSC-derived neurons.

Figure 2. B-27 Plus Neuronal Culture System enables superior cultures of primary rodent and induced pluripotent stem cell (iPSC)–derived neurons compared to B-27 Supplement and Neurobasal Medium. Cryopreserved neurons were cultured for 3–4 weeks in the listed media system. Neurons were immunostained at the indicated time points for the neuronal dendritic marker MAP2 (green) and the neuronal cell body marker HuC/D (red). Nuclei were counterstained with DAPI (blue).

Improves electrophysiological activity and maturity

B-27 Plus Neuronal Culture System is exhibits highly synchronized neuronal activity over time.

Figure 3. Improvement of electrophysiological activity compared to BrainPhys Neuronal Medium and SM1 Kit. Primary Rat Cortex Neurons plated on 48-well multi-electrode array (MEA) plates. Cells were cultured for 35 days in the listed media system following the suppliers’ recommended protocols. Spontaneous electrophysiological activity was recorded throughout with the Axion BioSystems™ Maestro™ MEA platform. Data shown are from one of four experiments, with each run showing similar trends. The B-27 Plus Neuronal Culture System enabled more consistent, stable, and highly synchronized spontaneous electrophysiological activity over time.

Enhanced neuronal maturation within healthy neuronal cell culture.

Figure 4. Enhancement of neuronal maturation compared to B-27 Supplement and Neurobasal Medium. Primary Rat Cortex Neurons at day 22 were stained for the dendritic marker MAP2 (red) and synapsin 1/2 to label presynaptic terminals (green). Neurons maintained in the B-27 Plus Neuronal Culture System had significantly higher synaptic-positive puncta.

B-27 Plus Neuronal Culture System promotes increased neural network activity.

Figure 5. The B-27 Plus Neuronal Culture System promotes increased neural activity. Primary rat cortex neurons were maintained for 35 days in vitro in B27 Plus system with half-medium changes every 2–3 days. Recordings were acquired 24 hours after media changes using Axion BioSystems Maestro Pro™ microelectrode array (MEA) platform. The B-27 Plus system exhibited neural coverage across the entire array (see activity map, left) and strong, synchronized network phenotype, as illustrated in the raster plot (activity map, right). (Data generated by Axion Biosystems.

Supports highest survival of neurons compared to alternative media systems

Figure 6. The B-27 Plus Neuronal Culture System achieves the highest neuronal survival in long-term cultures compared to other commercially available serum-free neuronal media systems. Primary Rat Cortical Neurons, Primary Rat Hippocampus Neurons, Primary Mouse Cortical Neurons and human iPSC-derived neurons that had been matured for 9 days before cryopreservation (HIP™ Neurons, MTI-GlobalStem) were thawed in classic Neurobasal Medium with B-27 Supplement. The neurons were plated onto poly-D-lysine–coated 96-well plates and maintained for 3–4 weeks in the B-27 Plus Neuronal Culture System and alternative serum-free supplemented media systems following the suppliers’ recommended protocols. Neuronal survival was quantitated by immunofluorescent labeling using Invitrogen HuC/HuD Monoclonal Antibody at 3 or 4 weeks. Data shown are from one of three experiments, with each run showing that B-27 Plus Neuronal Culture System enables the highest neuronal cell survival among the alternative systems.

Figure 7. Acceleration of neurite outgrowth compared to B-27 Supplement and Neurobasal Medium and alternative media systems. Primary Mouse Cortical Neurons were thawed in classic Neurobasal Medium with B-27 Supplement and plated onto poly-D-lysine–coated 96-well plates. Neurons were maintained for ~3 weeks in Neurobasal Medium with B-27 Supplement, Neurobasal Plus Medium with B-27 Plus Supplement, and other commercially available serum-free neuronal media systems following the suppliers’ recommended protocols. Neurite outgrowth was quantitated using differential interference contrast images taken at the time points specified. The B-27 Plus Neuronal Culture System significantly accelerates neurite outgrowth over the first few weeks compared to classic Neurobasal Medium with B-27 Supplement. Data shown are from one of three experiments, with each run showing after 1 week B-27 Plus Neuronal Culture System enabled the highest mean neurite length compared to all alternative systems.

Customer stories

Primary mouse hippocampal neruons (MHN) were cultivated over a period of 28 days with Neurobasal Plus Medium supplemented with B-27 Plus Supplement under standard conditions. The cells showed healthy morphology and no neurite fragmentation could be observed. All dendritic spine morphologies that can be found in vivo, could also be observed in cell culture. I would clearly recommend the Neurobasal Plus Medium and B-27 Plus Supplement for the long time cultivation of primary neurons.

— Susanne Zach, Boehringer Ingelheim, CNS Research

Susanne Zach
Professor Hideyuki Okano
The neurons in the B-27 Plus system appeared healthier, with longer and more complete neurites, as compared to the other media conditions. We are extremely happy with this new media system and plan on continuing to use it in our experiments as we believe that it will allow us to more efficiently and effectively study the differentiation and maturation of NSC and iPSC-derived neurons.

—  Professor Hideyuki Okano, MD, PhD, Keio University School of Medicine, Tokyo, Japan

With our primary rat cortical neural cultures, the B-27 Plus Neuronal Culture System produces robust neural activity across almost every electrode in MEA plates. The network bursting events are very regular, and extremely high intensity, making data analysis easy and straightforward when evaluating functional neural activity.

— Jim Ross, CTO at Axion BioSystems

Jim Ross
John Graef
B-27 Plus medium led to superior levels of spontaneous multielectrode array activity in our human iPSC-derived neurons as compared to our standard preparations with BrainPhys culture media. I found that B-27 Plus medium represents an excellent option for cell culture systems where identifying potential electrophysiological phenotypes based on spontaneous neuronal activity is desired.

— John Graef, PhD, Principal Scientist, Fulcrum Therapeutics

Recommended applications

B-27 Plus Supplement and Neurobasal Plus Medium come in the same formats and will work in exactly the same protocols as the current catalog products. The classic B-27 and Neurobasal products will continue to be offered; however, we recommend using the Plus versions and other combinations of medium and supplement in the following applications:

Application Recommended supplement(s) Recommended basal medium
Maintenance/maturation of prenatal/fetal primary neurons B-27 Plus Supplement Neurobasal Plus Medium
Differentiation and maintenance/maturation of stem cell–derived neurons B-27 Plus Supplement 
and
CultureOne Supplement
Neurobasal Plus Medium
Electrophysiology studies B-27 Plus Supplement Neurobasal Plus Medium
Maintenance/maturation of postnatal and adult brain neurons B-27 Plus Supplement Neurobasal-A Medium
Studies of oxidative stress/damage, apoptosis, or where free radical damage to neurons occurs B-27 Supplement minus Antioxidants Neurobasal Plus Medium

B-27 Plus System cost effectiveness

B-27 Plus Neuronal Culture System is cost effective compared to classic B27 supplement and Neurobasal Medium.

  B-27 Supplement and Neurobasal Medium B-27 Plus Neuronal Culture System
System price * $155 $182
System volume 500 mL 500 mL
Volume to maintain a 96-well plate of neurons for 2 weeks 60 mL 60 mL
Number of plates per system unit 8 8
Cost per plate * $19 $22
Relative neuronal survival (%B-27) 100% 150%
Total cost for same number of neurons * $19 $15
* Comparison is based on prices in US dollars.

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FAQs

Question Answer
What is the difference between B-27 Plus Supplement and classic B-27 Supplement? B-27 Plus Supplement is an optimized formulation of classic B-27. No small molecules or growth factors have been added.
What is the difference between Neurobasal Plus Medium and Neurobasal Medium? Key amino acids and buffering components have been optimized in Neurobasal Plus Medium. No small molecules or growth factors have been added.
How did you upgrade the manufacturing/QC process? We streamlined the manufacturing process, implemented more stringent acceptance criteria and enhanced QC assays for qualification of raw materials and release of finished product to enable better lot to lot consistency.
Can I re-freeze B-27 Plus into smaller aliquots for future use? Yes, we recommend aliquoting upon first thaw. We recommend no more than 2 total freeze/thaw cycles.
Can I use the B-27 Plus Neuronal Culture System for expansion of proliferating cells, such as neural stem cells? No, we do not recommend the B-27 Plus System for expanding proliferating cells. We recommend using StemPro NSC SFM, Neural Induction Medium Supplement or B-27 Supplement minus Vitamin A for growing proliferative cells, such as NSCs.
How long can I maintain my neurons in culture with the B-27 Plus System? We have maintained primary rat cortical neurons up to 8 weeks and rat hippocampal neurons up to 4 weeks. Mouse cortical neurons were maintained for 4 weeks. Human iPSC-derived neurons (HIP™ Neurons from MTI-GlobalStem) have been maintained for 5 weeks. All cells tested with the B-27 Plus System were significantly healthier than the classic products across all time points.
Do I have to use B-27 Plus with Neurobasal Plus? B-27 Plus Supplement and Neurobasal Plus Medium have been designed to work synergistically together for maximal performance. They can be used with other basal media or supplements if necessary.
How can I control the number of glial cells in my culture? Add CultureOne Supplement at day 0 with the B-27 Plus Neuronal Culture System to fully suppress both astrocytes and oligodendrocytes with no detrimental effect on neurons. Delaying the addition of CultureOne to later time points results in increasing levels of astrocytes. Refer to the CultureOne Supplement user guide for more information.
Can I use B-27 Plus Neuronal Culture System to culture non-neuronal cells? We have not tested in other cells, however B-27 Plus System was designed for culturing neuronal cells.
Which classic B-27 and Neurobasal products can I substitute B-27 Plus and Neurobasal Plus for? Substitute for the regular versions of B-27 Supplement and Neurobasal  Medium products (not specialty versions like minus Insulin, minus Vitamin-A or Neurobasal-A) in protocols used for:
- Maintenance of primary rat, mouse and human PSC-derived and fetal-derived neurons
- Differentiation of human PSC-derived and fetal-derived Neural Stem Cells (NSC) to neurons
How long can I use complete Neurobasal Plus supplemented media? Complete Neurobasal Plus Medium should be used within 4 weeks after supplementation with B-27 Plus Supplement.
Can thawed B-27 Plus Supplement be kept at 4C? Yes, B-27 Plus Supplement can be stored at 4C for up to 2 weeks post thaw.
What is the shelf-life of B-27 Plus Supplement and Neurobasal Plus Medium? One year
What do I do if my cells are not attaching or are clumped? In primary neuronal cultures this is typically a function of uneven coating or if neurons are plated at too high of a density. In NSC differentiation experiments, this could be due to over proliferation of neural progenitor cells. Differentiating with the B-27 Plus Neuronal Culture System and CultureOne Supplement can eliminate more than 75% of neural progenitor cells and the clumps they form. See CultureOne Supplement for more information.
What is HuC/D? HuC/D is an antibody specific to neuronal cell bodies. It can be used to quantitate neurons in culture.
How do I thaw B-27 Plus Supplement? Thaw at 4C overnight or at room temperature for 2 hours. We don't recommend thawing in a water bath or incubator.
What happens if I thaw B-27 Plus Supplement in a 37C water bath or incubator? This is not recommended.
How do I feed my cells with the B-27 Plus Neuronal Culture System? Perform half medium exchanges every 2-3 days post plating. Make sure that neurons are not completely exposed to air by not removing all of the medium during exchanges.

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