Vassanelli S, Bandiera L, Borgo M, Cellere G, Santoni L, Bersani C, Salamon M, Zaccolo M, Lorenzelli L, Girardi S, Maschietto M, Dal Maschio M, Paccagnella A
University of Padova, Department of Human Anatomy and Physiology, Section of Physiology, via Marzolo 3 - 35131, Padova, Italy.
N Biotechnol. 2008 Jun;25(1):55-67. doi: 10.1016/j.nbt.2008.03.002. Epub 2008 Apr 18.
Single-cell experiments represent the next frontier for biochemical and gene expression research. Although bulk-scale methods averaging populations of cells have been traditionally used to investigate cellular behavior, they mask individual cell features and can lead to misleading or insufficient biological results. We report on a single-cell electroporation microarray enabling the transfection of pre-selected individual cells at different sites within the same culture (space-resolved), at arbitrarily chosen time points and even sequentially to the same cells (time-resolved). Delivery of impermeant molecules by single-cell electroporation was first proven to be finely tunable by acting on the electroporation protocol and then optimized for transfection of nucleic acids into Chinese Hamster Ovary (CHO-K1) cells. We focused on DNA oligonucleotides (ODNs), short interfering RNAs (siRNAs), and DNA plasmid vectors, thus providing a versatile and easy-to-use platform for time-resolved gene expression experiments in single mammalian cells.
单细胞实验代表了生化和基因表达研究的下一个前沿领域。尽管传统上一直使用对细胞群体进行平均的大规模方法来研究细胞行为，但这些方法掩盖了单个细胞的特征，可能导致具有误导性或不充分的生物学结果。我们报道了一种单细胞电穿孔微阵列，它能够在同一培养物中的不同位点（空间分辨）、在任意选择的时间点，甚至对同一细胞进行连续转染（时间分辨），对预先选定的单个细胞进行转染。通过单细胞电穿孔递送非渗透性分子首先被证明可以通过作用于电穿孔方案进行精细调节，然后针对将核酸转染到中国仓鼠卵巢（CHO-K1）细胞中进行了优化。我们专注于DNA寡核苷酸（ODN）、小干扰RNA（siRNA）和DNA质粒载体，从而为单个哺乳动物细胞中的时间分辨基因表达实验提供了一个通用且易于使用的平台。
