( development and function we...)Biolistic Transfection of Organotypic Brain Slices

development and function, we use both neuronal cell lines (such as PC12 cells) and brain slice cultures. We have applied the Biolistic technology to transfect these preparations since both are difficult to transfect by conventional methods. Here, we provide information on optimizing the Biolistic parameters for transfection of dissociated cells and cultured brain slices (see Bio-Rad instruction manual; Arnold et al., 1994; Lo et al., 1994). Once optimized, Biolistic transformation is a reliable and efficient method for studying gene function in both cell lines and brain slice cultures.

Methods and Results
In general, particle bombardment consists of three steps: coating microprojectiles with DNA, drying them onto a macrocarrier disk and, finally, propelling them into target cells or tissue. The macrocarrier disk is accelerated with high-pressure helium into a stopping screen, which frees the microprojectiles to bombard the tissue. Cells penetrated by a microprojectile are likely to become successfully transfected.

The details of performing these procedures are described in the PDS-1000/He Biolistic instruction manual from Bio-Rad. We have made several modifications to these basic procedures that allow us to apply Biolistic bombardment to transfection of both neuronal cell lines and brain slice cultures (Figure 1).

Transfection of Neuronal Cell Lines
Culture Preparation
In accordance with requirements for each cell line, cells are plated on collagen-coated or untreated tissue culture plastic. To increase the number of successful transfections, the density of the cells should be high. Cells may be transfected as soon as they
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