Neural stem cells (NSC) are defined as cells that are capable of self-renewal and differentiation into neurons, astrocytes and oligodendrocytes. These cells can be isolated from either fetal or adult brain and cultured as neurospheres. Neurospheres can be cultured from NSC in a medium containing hormonal supplements and mitogens. Presence of epidermal growth factor (EGF) and basic fibroblast growth factor (b-FGF) is essential for culture of neurospheres. Initially, hormonal supplements such as N2 and B27 were used in the presence of growth factors for neurosphere culture. StemCell Technologies have come up with an NSC proliferation kit which contains a supplement that has a mixture of chemicals and hormones needed for optimal neurosphere growth. This is an excellent tool for neurosphere culture.
The Neurocult® NS-A Proliferation Kit comes with Neurocult NS-A basal medium and Neurocult NS-A proliferation supplements. The basal medium should be stored at 2-8°C and can be used with both the Neurocult NS-A proliferation and differentiation supplements. The basal medium is stable for six months and must be used with the proliferation supplements and cytokines for neurosphere growth. The proliferation supplements are supplied as a frozen mixture that needs to be aliquoted and stored at -20°C. Ten ml of supplements are to be added with 90 ml of basal medium along with EGF and b-FGF at 20 ng/ml and heparin at 2 µg/ml concentration to make the complete Neurocult proliferation medium. Although the manufacturer recommends preparation of complete medium with cytokines and heparin freshly before each use, our experience in the lab shows that the complete medium is stable at 4°C for about two weeks.
We have used the Neurocult NS-A Proliferation Kit in our laboratory for the last four years and have had excellent results. We were able to grow healthy neurospheres for over 4 months. However, it is crucial to dissociate the neurospheres and passage them before they become too large and necrotic to continue propagation. We have been able to generate and propagate neurospheres using the Neurocult NS-A proliferation kit from CD133 and nestin expressing fetal human brain derived neural stem cells (see figure below). The presence of all the required hormones and chemicals as a standard mixture in the supplements makes this kit very convenient and easy to use. However, the drawback of using this kit is that the scientist is unaware of the exact composition of the basalt medium as well as the supplements. This could pose a problem in experiments where knowledge of the medium composition is crucial.
In summary, the Neurocult NS-A proliferation kit is a very easy and convenient kit for growth of neurospheres. This would particularly be beneficial to labs that are just entering the neural stem cell arena and need an easy way to culture neurospheres. Additionally, the company also offers a neurosphere dissociation kit that could be used to dissociate neurospheres prior to passaging.
Figure 1: Floating neurospheres from fetal human brain derived neural stem cells grown using NeuroCult® NS-A Proliferation Kit (Human) from StemCell Technologies
Scientist
Children’s Research Center of Michigan
Children’s Hospital of Michigan