Zanacchi Francesca Cella, Manzo Carlo, Alvarez Angel S, Derr Nathan D, Garcia-Parajo Maria F, Lakadamyali Melike
ICFO-Institut de Ciencies Fotoniques, The Barcelona Institute of Science and Technology, Castelldefels, Spain.
Universitat de Vic - Universitat Central de Catalunya (UVic-UCC), Vic, Spain.
Nat Methods. 2017 Aug;14(8):789-792. doi: 10.1038/nmeth.4342. Epub 2017 Jun 26.
Single-molecule-based super-resolution microscopy offers researchers a unique opportunity to quantify protein copy number with nanoscale resolution. However, while fluorescent proteins have been characterized for quantitative imaging using calibration standards, similar calibration tools for immunofluorescence with small organic fluorophores are lacking. Here we show that DNA origami, in combination with GFP antibodies, is a versatile platform for calibrating fluorophore and antibody labeling efficiency to quantify protein copy number in cellular contexts using super-resolution microscopy.
基于单分子的超分辨率显微镜为研究人员提供了一个独特的机会，能够以纳米级分辨率对蛋白质拷贝数进行定量。然而，虽然荧光蛋白已通过校准标准进行了定量成像表征，但缺乏用于带有小有机荧光团的免疫荧光的类似校准工具。在这里，我们展示了DNA折纸与绿色荧光蛋白（GFP）抗体相结合，是一个多功能平台，可用于校准荧光团和抗体标记效率，以使用超分辨率显微镜在细胞环境中对蛋白质拷贝数进行定量。
