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of these cells with Fluoro-Gold injections into globus pallidus
externa and substantia nigra, respectively. We evaluated injec-
tion accuracy in each animal by blinded histological analysis
and our usual criterion is that a majority of transgene-express-
ing neurons must be located within the borders of the target
area bilaterally or the animal will be excluded from further
analysis. In some cases, missed injections may provide intrigu-
ing pilot data for subsequent studies or can be treated as ana-
tomical controls.
4
Notes
1. The use of a beveled needle for viral vector injections has been
found to enhance the quality of transgene expression, possibly
due to the increased surface area of the needle. If beveled nee-
dles are used, the bevel should be positioned toward the tar-
geted injection area. Slow injection speeds and needle
withdrawal also increase transgene expression, likely through
decreasing the chance of wicking the viral vector solution back
up the needle track.
2. When constructing a cell-specifi c viral vector, the viral pro-
moters should either be removed or the cell-specifi c promoter
should be placed in the opposite orientation relative to the
viral promoter to decrease the likelihood of leakage
expression.
3. During the surgical procedures, the virus should always be
kept on ice and away from light. The amount of viral vector
needed for an injection should be backfi lled into the injector
immediately prior to each injection.
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