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Proceedings Paper

SearchLight: a freely available web-based quantitative spectral analysis tool (Conference Presentation)
Author(s): Prashant Prabhat; Michael Peet; Turan Erdogan

Paper Abstract

In order to design a fluorescence experiment, typically the spectra of a fluorophore and of a filter set are overlaid on a single graph and the spectral overlap is evaluated intuitively. However, in a typical fluorescence imaging system the fluorophores and optical filters are not the only wavelength dependent variables - even the excitation light sources have been changing. For example, LED Light Engines may have a significantly different spectral response compared to the traditional metal-halide lamps. Therefore, for a more accurate assessment of fluorophore-to-filter-set compatibility, all sources of spectral variation should be taken into account simultaneously. Additionally, intuitive or qualitative evaluation of many spectra does not necessarily provide a realistic assessment of the system performance. “SearchLight” is a freely available web-based spectral plotting and analysis tool that can be used to address the need for accurate, quantitative spectral evaluation of fluorescence measurement systems. This tool is available at: Based on a detailed mathematical framework [1], SearchLight calculates signal, noise, and signal-to-noise ratio for multiple combinations of fluorophores, filter sets, light sources and detectors. SearchLight allows for qualitative and quantitative evaluation of the compatibility of filter sets with fluorophores, analysis of bleed-through, identification of optimized spectral edge locations for a set of filters under specific experimental conditions, and guidance regarding labeling protocols in multiplexing imaging assays. Entire SearchLight sessions can be shared with colleagues and collaborators and saved for future reference. [1] Anderson, N., Prabhat, P. and Erdogan, T., Spectral Modeling in Fluorescence Microscopy, (2010).

Paper Details

Date Published: 27 April 2016
PDF: 1 pages
Proc. SPIE 9712, Multiphoton Microscopy in the Biomedical Sciences XVI, 971210 (27 April 2016); doi: 10.1117/12.2219999
Show Author Affiliations
Prashant Prabhat, Semrock, Inc. (United States)
Michael Peet, Semrock, Inc. (United States)
Turan Erdogan, Semrock, Inc. (United States)

Published in SPIE Proceedings Vol. 9712:
Multiphoton Microscopy in the Biomedical Sciences XVI
Ammasi Periasamy; Peter T. C. So; Karsten König, Editor(s)

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