In vitro culturing of primary neurons is a mainstay of neurobiological research. Many of these culture paradigms have taken advantage of defined culture media rather than serum additives that contain undefined survival factors to facilitate experimental manipulations and interpretation of the results. To culture neurons in the absence of serum, defined supplements such as B27 are now widely used. However, commercially available supplements exhibit large variability in their capabilities to support neurons in culture. We re-optimized and modified earlier published formulations of B27 using 21 different ingredients (NS21). NS21 supports neuronal cultures of high quality as manifested by their morphological characteristics, formation of synapses, and postsynaptic responses. Much of the variability in the quality of B27/NS21 was due to variability in the quality of different sources of bovine serum albumin. Furthermore, we found that holo-transferrin used in NS21 is preferable over apo-transferrin used in B27 for the quality of neuronal cultures.

Original languageEnglish
Pages (from-to)239-247
Number of pages9
JournalJournal of Neuroscience Methods
Issue number2
StatePublished - Jun 30 2008


  • B27 supplement
  • DRG
  • Dorsal root ganglion cells
  • Hippocampal neurons
  • NS21 supplement
  • Neuronal cultures
  • Primary hippocampal cultures
  • RGC
  • Retinal ganglion cells


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