Delivery

TABLE (Is this from Wikipedia?)

Note that the method of delivery determines the dose of DNA required to raise an effective immune response. For example, each saline IM injection requires 100μg – 200μg of pDNA whilst gene gun delivery needs just around a thousandth the amount of pDNA to trigger the same effective immune response.

The two most popular approaches are injection of DNA in saline, using a standard hypodermic needle, and a gene gun delivery. In gene gun delivery, pDNA that has been adsorbed onto gold or tungsten nanoparticles are ballistically accelerated into the target cells, basically bombarding them, using compressed helium as the accelerant[1]. Whilst uptake is efficient using the gene delivery method, the opposite is the case for saline injection.

Optimisation

In order to optimise the potency of DNA vaccines in saline injections, electroporation is often used to increase host cell transfection rate[2]. This can be carried out by simply using hypertonic solutions of saline or sucrose. An alternative method (when injecting intramuscularly) is using a myotoxin such as bupivacaine to temporarily damage muscle fibres.

PICTURE

Factors influencing the efficiency of gene electrotransfer:

• Temperature
• Parameters of electric pulses
• pDNA concentration
• host cell size
• ability of host cells to express transfected genes (e.g. muscle tissue is more able to do so[1])

Our project E.coili makes a contribution here by altering the plasmid itself to increase expression as well as devising formulations to augment the stability of the plasmid as well as enabling the alternate delivery of pDNA mucosally due to supercoiling.