Bio-Rad Helios® Gene Gun System - Manual (Page 8)

Prior to transfection, the plasmid DNA must be attached to the gold particles. This is accom-
plished by precipitation of the DNA from solution in the presence of gold microcarriers and the
polycation spermidine by the addition of CaCl
2
. The particles are then washed extensively with ethanol
to remove the water and resuspended in ethanol. Using the Tubing Prep Station, the DNA/microcar-
rier solution is coated onto the inner wall of Gold-Coat tubing and dried. The tubing is then cut into 0.5"
length cartridges using the Tubing Cutter. These cartridges, when inserted into the cartridge holder of
the Helios Gene Gun are the source of the DNA which enters the target cells by the helium discharge.
The Helios Gene Gun employs a high velocity stream of helium to accelerate gold particles
coated with plasmids or RNA to velocities sufficient to penetrate and transform cells, both in vitro
and in vivo (Figure 2). The discharge is initiated by pressing the trigger buttons which activates the
main valve, causing helium to travel down the bore of the particle delivery device. When the heli-
um enters one of the bores of the cylinder containing the cartridge, the gold particles on the inside
of the tubing are pulled from the surface, become entrained in the helium stream, and begin to
pick up speed. Immediately past the acceleration channel, the barrel begins to open as a cone. The
slope of the cone causes the gas to be pulled outward, a process known as the Coanda effect (Reba,
1966), expanding the high pressure jet into a less destructive low velocity pulse, while the gold par-
ticles maintain a high velocity. The expansion also helps spread the microcarriers from their original
1/16" diameter to an area approximately 1/2" in diameter at the target site.
Fig. 2. How the samples are delivered. Helium gas is pulsed through the cartridge loaded with DNA-coat-
ed microcarriers. This pulse sweeps the microcarriers from the inside wall of the cartridge. As the microcarriers
enter the barrel liner they pick up speed in the acceleration channel then spread out as they travel down the
barrel; the increased cross-sectional area of the barrel from the acceleration chamber to the spacer also
moderates the helium shock wave so it is less intense when it reaches the target cells. The O-rings on each
side of the cartridge holder direct the flow of helium through the cartridge and the acceleration channel. The
spacer maintains optimal target distance and permits venting of the helium gas away from the target.
2.4 Requirements for System Operation
Selecting a Site for Operation
The Helios Gene Gun is a portable particle bombardment device. The range of its use is
limited by its requirement for a supply of pressurized helium and the 6 foot length of pres-
surized helium hose. When using the Gene Gun, only a small area is needed for setting down
the gun during an experiment, for loading the cartridges into the cartridge holders and exchang-
ing cartridge holders during experiments. In addition, a clean and dry area is needed for
working with the tissue samples.
4