Nucleofector device for the efficient transfer of genetic material into hard-to-transfect mammalian cells

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Macrophages (MP) and dendritic cells (DC) are specialized cells of the immune system that play an essential role in protecting us from disease by directly eliminating invading pathogens and relaying the danger signal to lymphocytes. Because of these properties, MP and DC are also major targets of infection by several viruses and bacteria. Two such pathogens are human Cytomegalovirus (CMV) and Burkholderia cepacia, which are major causes of disease in immunocompromised and in cystic fibrosis (CF) patients, respectively. To fight against these and other pathogens, MP and DC produce several inflammatory mediators, whose uncontrolled release can potentially trigger the onset of diseases such as rheumatoid arthritis. Understanding the molecular mechanisms regulating MP and DC activation as well as the relationship between these cells and specific pathogens is vital for the development of new anti-viral, anti-microbial and anti-inflammatory therapies. In addition, the possibility to biotechnologically manipulate these cells can lead to their use in the eradication of cancers, the stimulation of vaccine efficacy, and the induction of immune tolerance. Primary MP and DC, however, are particularly difficult to manipulate because their ability to accept foreign DNA is extremely low. This limitation strongly compromises progress in the applicants' research, ultimately slowing the pace at which the above mentioned novel therapies are designed. This proposal is for the acquisition of a Nucleofector 96-well Shuttle System, a device that has been specifically conceived to deliver foreign DNA into these cells with high efficiency. This instrument will become a central piece of equipment for trainees in the laboratories of the three applicants and will allow the rapid generation of highly valuable scientific data which would otherwise be unattainable. To our knowledge no other equipment with similar capabilities exists on campus.