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

巨噬细胞(MP)和树突状细胞(DC)是免疫系统中的特殊细胞，通过直接清除入侵的病原体和将危险信号传递给淋巴细胞，在保护我们免受疾病侵袭方面发挥着重要作用。由于这些特性，MP和DC也是几种病毒和细菌感染的主要目标。两种这样的病原体是人类巨细胞病毒(CMV)和洋葱伯克霍尔德氏菌，分别是免疫功能低下和囊性纤维化(CF)患者的主要致病原因。为了对抗这些和其他病原体，MP和DC会产生几种炎症介质，它们的不受控制的释放可能会引发类风湿性关节炎等疾病的发病。了解调节MP和DC激活的分子机制以及这些细胞与特定病原体之间的关系对于开发新的抗病毒、抗微生物和抗炎疗法至关重要。此外，对这些细胞进行生物技术操作的可能性可以导致它们用于根除癌症、刺激疫苗效力和诱导免疫耐受。然而，初级MP和DC尤其难以操纵，因为它们接受外来DNA的能力极低。这一限制严重影响了申请者的研究进展，最终减缓了上述新疗法的设计速度。这项提议是为了获得一个96孔核探测器穿梭系统，这是一种专门设计的设备，可以高效地将外来DNA输送到这些细胞中。该仪器将成为三个申请者实验室中受训人员的核心设备，并将能够迅速产生高价值的科学数据，否则这些数据是无法获得的。据我们所知，校园里没有其他具有类似能力的设备。

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.