Illumination Power, Stability, and Linearity Measurements for Confocal and Widefield Microscopes

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dc.catalogadorjwg
dc.contributor.authorGaudreault, Nathalie
dc.contributor.authorAli, Rizwan
dc.contributor.authorAvilov, Sergiy V.
dc.contributor.authorBagley, Steve
dc.contributor.authorBammann, Rodrigo R.
dc.contributor.authorBarachati, Fabio
dc.contributor.authorBertocchi, Cristina
dc.contributor.authorBoehm, Ulrike
dc.contributor.authorBosch, Manel
dc.contributor.authorBrideau, Craig
dc.contributor.authorCarvalho, M. T.
dc.contributor.authorColarusso, Pina
dc.contributor.authorDuwé, Sam
dc.contributor.authorGuilbert, Thomas
dc.contributor.authorKirchner, Marcel
dc.contributor.authorAydemir-Koksoy, Ayse Aslihan
dc.contributor.authorKrens, Gabriel
dc.contributor.authorLaude, Alex
dc.contributor.authorLedue, Jeffrey M.
dc.contributor.authorLiu, Penghuan
dc.date.accessioned2024-01-31T14:06:23Z
dc.date.available2024-01-31T14:06:23Z
dc.date.issued2023
dc.description.abstractTo obtain accurate, reproducible, and interpretable data when conducting imaging experiments, it is critical to consider external factors affecting data acquisition at various steps of the experimental workflow. Illumination power and stability represent two critical factors, especially when comparing fluorescence intensities between images during a time-lapse experiment or experiments performed at different times or on different microscopes. The fluorescence signal can be generated by different types of light sources. These light sources and their coupling elements (e.g., fibers) can display varying performances over time as they age, move, or as environmental conditions change. Unfortunately, microscope users can often only set illumination power as a percentage of its maximal output and may, therefore, not be aware of potential performance changes. It is important to recognize that a set percentage will not always yield the same illumination power in Watts at the objective over the course of an experiment, not to mention between days or systems. This means that selecting for example 10% output may lead to different experimental results over time or even between two microscopes of the same model. In addition to illumination stability, working within the linear range of the illumination power allows to adjust accurately the illumination power absolute value (in mW) using fraction (or %) of its maximal value through the imaging software. If you are responsible for system maintenance, routinely measuring the illumination power, stability, and linearity over time can help you detect issues that affect the integrity of the system and thus the reproducibility of an experiment. This protocol describes how to measure the stability and linearity of the illumination power using calibrated external power sensors. This protocol is at the moment intended for confocal systems (raster scanning and spinning disks), and widefield systems. It represents the collective experience of over 50 imaging scientists. Measurements made by our working group with this protocol are available in a public database, which will be updated with further contributions from the community.
dc.fuente.origenORCID
dc.identifier.doi10.17504/protocols.io.5jyl853ndl2w/v2
dc.identifier.urihttps://www.protocols.io/view/illumination-power-stability-and-linearity-measure-b68prhvn
dc.identifier.urihttps://repositorio.uc.cl/handle/11534/81112
dc.information.autorucFacultad de Ciencias Biológicas; Bertocchi Cristina; 0000-0003-0907-1318; 1078032
dc.language.isoen
dc.nota.accesoContenido completo
dc.rightsacceso restringido
dc.subject.ddc610
dc.subject.deweyMedicina y saludes_ES
dc.titleIllumination Power, Stability, and Linearity Measurements for Confocal and Widefield Microscopes
dc.typeartículo
sipa.codpersvinculados1078032
sipa.trazabilidadORCID;2024-01-15
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