2.1. Isolation and cultivation of primary neuronal cells
Protocols for the cultivation of honey bee neuronal cells were developed about twenty years ago (Gascuel et al., 1991; Kreissl and Bicker, 1992; Devaud et al., 1994; Gascuel et al., 1994;). The original purpose at that time was to complement in vivo studies on the insect olfactory system with data from in vitro cell culture experiments. Unfortunately, these protocols did not find their way from bee neuroscience into bee pathology until recently, when they were adopted for cultivation of several cell types originating from pupal or adult brain and gut (Möckel et al., 2009; Poppinga et al., 2012). Although these primary cells proved to be useful, they have their limitations. Most primary cells stay viable only for a limited time period or, if it is possible to split the culture, they can be passaged several times only as a non-permanent cell line before they die. During these passages most cells change their characteristics to adapt to the artificial environment, which sometimes creates problems with reproducibility of results.