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This page is intended to be a repository of useful information for STORM sample preparation and imaging.
Any dye which can be switched from an off state to an on state can be used for super-resolution imaging by localization microscopy. This has led to a large number of different experimental designs in the literature. Here we highlight a few commonly used approaches.
In order to collect good STORM imaging data sample prep is key. Below are important aspects to optimize and consider.
Dani A, Huang B, Bergan J, Dulac C, Zhuang X. Superresolution imaging of chemical synapses in the brain. Neuron. 2010 Dec 9;68(5):843-56. Describes using three color STORM imaging to localize proteins within synapses in 10 um brain sections.
Lana Lau, Yin Loon Lee, Steffen J. Sahl, Tim Stearns, and W. E. Moerner. STED Microscopy with Optimized Labeling Density Reveals 9-Fold Arrangement of a Centriole Protein. Biophysical Journal. 2012 June 102:2926–2935 This paper has a nice test of the effect of antibody labeling density on image resolution using STED microscopy. While they study the effect of labeling density on STED imaging, I suspect much of what they find is relevant to STORM and SIM imaging as well.
Ries J, Kaplan C, Platonova E, Eghlidi H, Ewers H. A simple, versatile method for GFP-based super-resolution microscopy via nanobodies. Nat Methods. 2012 Apr 29 This paper demonstrates the use of a single chain antibody (nanobody) against GFP for STORM imaging of GFP tagged proteins, by binding an Alexa 647 labeled nanobody to GFP-tagged proteins. The nanobody is commercially available from here: http://www.chromotek.com/home/
Lew MD, Lee SF, Ptacin JL, Lee MK, Twieg RJ, Shapiro L, Moerner WE. Three-dimensional superresolution colocalization of intracellular protein superstructures and the cell surface in live Caulobacter crescentus. Proc Natl Acad Sci U S A. 2011 Nov 15;108(46):E1102-10. A localization microscopy experiment combining single molecule blinking of eYFP and localization of Nile Red by random insertion of the dye into the membrane at very low concentration.
Jones SA, Shim SH, He J, Zhuang X. Fast, three-dimensional super-resolution imaging of live cells. Nat Methods. 2011 Jun;8(6):499-508. This paper demonstrates fast STORM imaging of live cells; it also compares the performance of a number of photoswitchable dyes and mEos2 and tdEos.
Shroff H, Galbraith CG, Galbraith JA, White H, Gillette J, Olenych S, Davidson MW, Betzig E. Dual-color superresolution imaging of genetically expressed probes within individual adhesion complexes. Proc Natl Acad Sci U S A. 2007 Dec 18;104(51):20308-13. This paper describes a number of methods for two-color PALM imaging using genetically encoded fluorescent proteins.
Dempsey GT, Vaughan JC, Chen KH, Bates M, Zhuang X. Evaluation of fluorophores for optimal performance in localization-based super-resolution imaging. Nat Methods. 2011 Nov 6;8(12):1027-36. This paper directly compares the performance of 26 different dyes for single dye localization microscopy (dSTORM). An excellent resource for choosing dyes for multicolor imaging.
Dempsey, Wang, and Zhuang. Fluorescence Imaging at Sub-Diffraction-Limit Resolution with Stochastic Optical Reconstruction Microscopy. In Handbook of Single-Molecule Biophysics, Hinterdorfer and Van Oijen, eds.