Culturing Nerve Cells
A do-it-yourself manual for culturing nerve cells, complete with recipes and protocols.
Because neurons and glia in culture are remarkably similar to those in situ, culture systems make it possible to identify significant cell interactions and to elucidate their mechanisms. This book is in many ways a do-it-yourself manual for culturing nerve cells, complete with recipes and protocols. But it also provides an understanding of the principles behind the protocols. In effect the contributors invite you into their labs and provide much of the information you would obtain from such a visit.The authors of the introductory chapters present the nuts-and-bolts principles of growing nerve cells. The authors of the following chapters discuss the culturing of specific cell types. They explain how their experimental goals have shaped their particular cell culture approach and the advantages and disadvantages of the cell culture systems they have developed. They provide detailed protocols and describe their cultures in practical terms, from when the cells are first plated through the various phases of their development.
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To overcome the limitations in efficiency with physical methods, many
neuroscientists have turned to viral methods. These methods make use of the
natural means viruses have evolved for ensuring that their DNA reaches the
nucleus of ...
An advantage of viral vectors is that they can be used to transfect neurons in vivo
with fairly high efficiency, and in fact most have been developed by labs
interested in potential applications in gene therapy. All these viral vectors can
Table 4.1 Wiral Wectors for Neuron Transfection HELPER MAXIMUM TITERS*
EXPRESSION CONSTRUCTION VIRAL VECTOR DEFECTIVE REQUIRED
GENOME INSERT (pfu/ml) TIME TIME R600mbinant Yes No 150 kb DNA >30 kb
A novel method that is not quite efficient enough for routine use but that seems
promising is the production of recombinant adenovirus deleted of all viral genes
that are propagated in the presence of an E1-deleted helper virus (Fisher et al., ...
Thus, they are the least toxic of viral vectors. The method of construction involves
cotransfection of a plasmid containing cis-acting replication sequences and the
expression cassette with a nonpackaging complementing AAV plasmid and then
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