Simultaneous spectral illumination of microplates for high-throughput optogenetics and photobiology

被引：0

作者：

Vogt, Arend ^{[1
,2
,3
,4
]}

Paulat, Raik ^{[1
,2
,3
,6
]}

Parthier, Daniel ^{[2
,3
,4
]}

Just, Verena ^{[1
,3
]}

Szczepek, Michal ^{[2
,3
,7
]}

Scheerer, Patrick ^{[2
,3
,7
]}

Xu, Qianzhao ^{[8
]}

Moeglich, Andreas ^{[8
]}

Schmitz, Dietmar ^{[2
,3
,4
]}

Rost, Benjamin R. ^{[2
,3
,4
,5
]}

Wenger, Nikolaus ^{[1
,2
,3
]}

机构：

[1] Charite Univ Med Berlin, Dept Neurol Expt Neurol, Translat Neuromodulat Grp, D-10117 Berlin, Germany

[2] Free Univ Berlin, D-10117 Berlin, Germany

[3] Humboldt Univ, D-10117 Berlin, Germany

[4] Charite Univ Med Berlin, Neurosci Res Ctr, D-10117 Berlin, Germany

[5] German Ctr Neurodegenerat Dis DZNE, D-10117 Berlin, Germany

[6] HTW Berlin Univ Appl Sci, Fac Energy & Informat, D-10318 Berlin, Germany

[7] Charite Univ Med Berlin, Inst Med Phys & Biophys, Grp Struct Biol Cellular Signaling, D-10117 Berlin, Germany

[8] Univ Bayreuth, Dept Biochem, D-95447 Bayreuth, Germany

来源：

BIOLOGICAL CHEMISTRY | 2024年 / 405卷 / 11-12期

关键词：

action spectra; cyclic mononucleotides; GPCR; photoactivated nucleotidyl cyclases; rhodopsin; signal transduction; CRYSTAL-STRUCTURE; ADENYLYL-CYCLASE; BOVINE RHODOPSIN; BINDING-PROTEIN; LIGHT; MECHANISM; DESIGN; OPSIN; GENE; CAMP;

D O I：

10.1515/hsz-2023-0205

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

The biophysical characterization and engineering of optogenetic tools and photobiological systems has been hampered by the lack of efficient methods for spectral illumination of microplates for high-throughput analysis of action spectra. Current methods to determine action spectra only allow the sequential spectral illumination of individual wells. Here we present the open-source RainbowCap-system, which combines LEDs and optical filters in a standard 96-well microplate format for simultaneous and spectrally defined illumination. The RainbowCap provides equal photon flux for each wavelength, with the output of the LEDs narrowed by optical bandpass filters. We validated the RainbowCap for photoactivatable G protein-coupled receptors (opto-GPCRs) and enzymes for the control of intracellular downstream signaling. The simultaneous, spectrally defined illumination provides minimal interruption during time-series measurements, while resolving 10 nm differences in the action spectra of optogenetic proteins under identical experimental conditions. The RainbowCap is also suitable for studying the spectral dependence of light-regulated gene expression in bacteria, which requires illumination over several hours. In summary, the RainbowCap provides high-throughput spectral illumination of microplates, while its modular, customizable design allows easy adaptation to a wide range of optogenetic and photobiological applications.

引用

页码：751 / 763

页数：13