BIO RAD Helios Gene Gun System, Инструкция по эксплуатации

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Section 10 Appendices
48 | Helios Gene Gun System
Discharge into Parafilm
Materials
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Parafilm Laboratory Film (Bemis Company, Inc., Neenah, WI)
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Glass plate
Procedure
1. Cut a piece of Parafilm from the roll; a 1" length is required for each cartridge to be assayed.
2. Smooth and press the Parafilm laboratory film (waxy surface down) onto the glass plate. Fold the edges
around the sides then peel away the protective paper.
3. Connect the helium regulator, helium hose, and Helios Gene Gun as described in Connecting the Helios
Gene Gun to a Helium Source, in Section 4.
4. Pressurize the system as described in Particle Delivery Using the Helios Gene Gun, in Section 5.
5. Discharge a cartridge at the test psi into a strip of Parafilm Laboratory Film. At this point, one can
roughly determine the quality of the microcarrier preps by examining the pattern of gold particles. When
examining the resulting pattern, it should appear circular with a dark center (Figure 25). The discharged
cartridges should be completely empty.
Spot darkness Proportional to particle penetration
Spot width Proportional to particle spread
Spot shape Uniformity of shot
Discharge into 3% Water Agar
Materials
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Slides, coverslips, and mounting solution
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Microscope with 10x eyepiece micrometer
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3% water agar in 60 mm petri dishes
Procedure
1. Discharge a cartridge at the test psi into 3% water agar. At this point, one can roughly determine
the quality of the microcarrier preps by examining the pattern of gold particles. When examining the
resulting pattern, it should appear circular with a dark center. The discharged cartridges should be
completely empty.
Additionally, one can also examine microcarrier depth penetration into the water agar microscopically.
Cut a thin slice approximately 0.5 cm long through the center of the agar target. Mount the slice onto a
slide with mounting solution and a coverslip and analyze for microcarrier depth and concentration.
2. A band of microcarriers will be visible in the agar with increasing particle sizes at increased depths.
The researcher will be able to determine areas of high, medium, and low microcarrier density in each
slice (Figure 26).