@article {65972,
	title = {High throughput instrument to screen fluorescent proteins under two-photon excitation.},
	journal = {Biomedical Optics Express},
	volume = {11},
	year = {2020},
	month = {2020 Dec 01},
	pages = {7192-7203},
	abstract = {<p>Two-photon microscopy together with fluorescent proteins and fluorescent protein-based biosensors are commonly used tools in neuroscience. To enhance their experimental scope, it is important to optimize fluorescent proteins for two-photon excitation. Directed evolution of fluorescent proteins under one-photon excitation is common, but many one-photon properties do not correlate with two-photon properties. A simple system for expressing fluorescent protein mutants is colonies on an agar plate. The small focal volume of two-photon excitation makes creating a high throughput screen in this system a challenge for a conventional point-scanning approach. We present an instrument and accompanying software that solves this challenge by selectively scanning each colony based on a colony map captured under one-photon excitation. This instrument, called the GIZMO, can measure the two-photon excited fluorescence of 10,000 colonies in 7 hours. We show that the GIZMO can be used to evolve a fluorescent protein under two-photon excitation.</p>
},
	issn = {2156-7085},
	doi = {10.1364/BOE.409353},
	author = {Molina, Rosana S and King, Jonathan and Franklin, Jacob and Clack, Nathan and McRaven, Christopher and Goncharov, Vasily and Flickinger, Daniel and Svoboda, Karel and Drobizhev, Mikhail and Hughes, Thomas E}
}
