I am testing the differences between serum-free media (neurobasal + B27) and serum media (HBSS + horse serum) on the viability of organotypic hippocampal slice cultures. For slice culture, as opposed to dissociated cell culture, I know that the media components for the dissection and culturing are different than that of dissociated culture to maintain the integrity of the tissue rather than wanting the cells to come apart from one another. Can you help me understand the differences in the required media components between the two, and what those components do, and the role of serum in each technique? Thank you very much.
Dear Chloe,
You do not mention the age of the animals from which you obtain the hippocampal slices. This may make a difference in your results.
Basically, the difference between serum-containing media and serum-free media is that serum is a bit of a "black box" since it contains lots of goodies that cells need. However, there are batch-to-batch differences, and even source-to-source differences. Serum-free media has defined components, the mix of which has been shown to keep the cell type of interest alive and healthy. This eliminates the need for batch testing of serum lots each time you need to order new serum, as well as overcoming serum shortages which occur. It also makes the interpretation of results of pharmacological studies easier.
The basal media that you choose also plays a role in this, since they all have different components that may be helpful, or not. For instance, Neurobasal media was developed to overcome some of the osmolarity and amino acid concentration problems created by DME and DME/F12 mixtures for neuronal survival. So, in your example, HBSS may be a little too simple with only horse serum supplementation to support the survival of the many cell types (not just neurons) in your hippocampal slice.
A good explanation of this can be found in Chapter 19, written by Paul J. Price and Gregory J. Brewer, of Protocols for Neural Cell Culture, 3rd Edition, 2001. Ed.: S. Federoff and A. Richardson. A link to an online copy is here:
http://www.bioon.com.cn/ewebeditor/uploadfile/201012/20101221133346840.pdf
Many people that I know use either the Romijn or Annis medias for slice cultures, which are much more complex than just Neurobasal and B27. The information is found in these two publications:
Romijn HJ, de Jong BM, Ruijter JM. A procedure for culturing rat neocortex
explants in a serum-free nutrient medium. J Neurosci Methods. 1988
Feb;23(1):75-83. PubMed PMID: 3347091.
Annis CM, Edmond J, Robertson RT. A chemically-defined medium for organotypic
slice cultures. J Neurosci Methods. 1990 Apr;32(1):63-70. PubMed PMID: 1970842.
As for the differences between dissection media and growth media, the first maintains the health of the tissue while you are cutting the slices, while growth media keeps everything happy for a long time in the culture dish. There are also lots of different dissection media. Usually they have components that block neuronal excitotoxicity, extra glucose to help with cell survival, and maintain pH and ionic gradients of the cells while they are in air on the bench (rather than CO2 in the incubator). Use whatever most protocols suggest for your tissue of interest.
Good Luck!
Jill