High-Efficiency Biolistic Device for Brain Transfection

基于粒子介导的基因转移（或“生物弹击”）的设备，用于识别和验证候选治疗药物和靶标。新的基因组学和蛋白质组学技术的巨大步伐正在推进，并产生关于正常和病理大脑的新信息，这已经在转化神经科学中产生了一个主要瓶颈，即生理学验证新的候选治疗靶点的能力，从而在药物化学中建立靶点驱动的努力。在这种情况下，一个主要的技术限速步骤一直是难以操纵的基因和蛋白质的表达在神经疾病特异性实验系统的基础上真正的，有丝分裂后的神经元，从原代培养组织外植体模型。 在初步的实验中，我们已经证明了一种基于 粒子介导的基因转移或生物射弹技术可用于在活脑切片中创建的疾病特异性模型中以中等通量水平筛选基于DMA的文库和化学文库。我们建议开发一种新的基于粒子加速的技术，以提高转染效率和通量10倍或更多。这种改进且价格实惠的生物枪装置将使标准规模的实验室小组能够利用现有的大量基因组和蛋白质组信息，在神经细胞和组织中进行高内容、功能筛选和验证实验。研究计划有三个具体目标： 具体目标1：研制一种新型高效的粒子加速转染装置。 具体目标2：确定新设备的射击参数的可行范围。 具体目标3：评价新生物射弹器械相对于现有技术的效率和有效性。

device, based on particle-mediated gene transfer (or 'biolistics'), for the identification and validation of candidate therapeutic drugs and targets. The tremendous pace at which new genomic and proteomic technologies are advancing and generating new information regarding the normal and pathological brain has created a major bottleneck in translational neuroscience, namely, the ability to physiologically validate new candidate therapeutic targets upon which to base target-driven efforts in medicinal chemistry. In this context, a major technological rate-limiting step has been the difficulty of manipulating the expression of genes and proteins in neural disease-specific experimental systems based on bona fide, postmitotic neurons, ranging from primary cultures to tissue explant models. In preliminary experiments, we have demonstrated that a neural transfection approach based on particle-mediated gene transfer, or biolistics, can be used to screen DMA-based and chemical libraries at medium-throughput levels in disease-specific models created in living brain slices. We propose here to develop a new particle acceleration-based technology to increase transfection efficiency and throughput by 10-fold or more. Such an improved and affordable biolistic device would allow laboratory groups of standard size to embark upon high-content, functional screening and validation experiments in neural cells and tissues using the mass of genomic and proteomic information now available. The Research Plan has 3 specific aims: Specific Aim1: To develop a new and highly efficient particle-acceleration transfection device. Specific Aim 2: To determine workable ranges of shooting parameters for the new device. Specific Aim 3: To evaluate the efficiency and effectiveness of the new biolistic device with respect to existing technology.