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arXiv · 1002.1980

Single-image measurements of monochromatic subdiffraction dimolecular separations

Shawn H. DeCenzo·Michael C. DeSantis·Y. M. Wang
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Abstract

Measuring subdiffraction separations between single fluorescent particles is important for biological, nano-, and medical-technology studies. Major challenges include (i) measuring changing molecular separations with high temporal resolution while (ii) using identical fluorescent labels. Here we report a method that measures subdiffraction separations between two identical fluorophores by using a single image of milliseconds exposure time and a standard single-molecule fluorescent imaging setup. The fluorophores do not need to be bleached and the separations can be measured down to 40 nm with nanometer precision. The method is called single-molecule image deconvolution -- SMID, and in this article it measures the standard deviation (SD) of Gaussian-approximated combined fluorescent intensity profiles of the two subdiffraction-separated fluorophores. This study enables measurements of (i) subdiffraction dimolecular separations using a single image, lifting the temporal resolution of seconds to milliseconds, while (ii) using identical fluorophores. The single-image nature of this dimer separation study makes it a single-image molecular analysis (SIMA) study.

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Shawn H. DeCenzo, Michael C. DeSantis, Y. M. Wang. 2010-08-31. Single-image measurements of monochromatic subdiffraction dimolecular separations. https://doi.org/10.1016/j.bpj.2009.12.3203

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