ACT-LABS gun system Instruction Manual download pdf (Page 39)

Table 4. Suggested starting conditions for

in vitro

transformation

of tissue culture cells using the Helios Gene Gun.

Parameter Conditions

Helium pressure: 50–200 psi

PVP: 0–0.015 mg/ml

MLQ: 0.125–0.5 mg/tube

Microcarriers: 1.0 or 1.6 µ gold

DLR: 0.5–2.5 µg DNA/mg gold (0.06–1.25 µg DNA/cartridge)

7.3 Parameters for

in vivo

Delivery

The following are suggested starting conditions for optimizing DNA delivery to the skin of

various species. Each laboratory should determine the optimum parameters for their particular

application (see Section 7.1 and Table 5). Bombardment conditions are those used in references

cited in Section 10.8 using the Accell Gene Gun. Use this information as a starting point when

optimizing bombardment conditions with the Helios Gene Gun. Lower helium pressures should

be tested early in the optimization process.

Table 5. Examples of parameters for delivery into skin of various

species

Species Site MLQ(mg/target) DLR(µg/mg) PVP(mg/ml) psi

Dog

Dorsum 0.5 1–2 0.1 300

Monkey

7, 8, 9

Abdomen 0.25 2 0–0.05 300–400

Mouse

17, 19

Abdomen* 0.25–0.5 1–2.5 0–0.1 300–500

Pig

7, 8, 10

Inner thigh 0.25–0.5 0.5–2.5 0.05 500

(clipping not

necessary)

Rabbit

Back 0.5 1–2 None 350–400

* Do not use a depilatory agent to remove the fur (Section 6.3, step 3)

Section 8

Troubleshooting

8.1 DNA/Microcarrier Preparation

Problem Microcarriers agglomerate after coating with DNA.

Possible solutions Lower DNA Loading Rate.

8.2 Cartridge Preparation

Problem Gold does not spread evenly in the Gold-Coat tubing (rings,

clumps, uncoated sections, streaks).

Possible solutions Eliminate any potential sources of water in the tube and in the

final DNA/Microcarrier preparation.

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