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Influence of ClearT and ClearT2 Agitation Conditions in the Fluorescence Imaging of 3D Spheroids

2020-12-29Journal article Open access
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dc.contributor.authorSilva, Daniel N.
dc.contributor.authorCosta, Elisabete
dc.contributor.authorRodrigues, Ana Carolina Félix
dc.contributor.authorDiogo, Duarte de Melo
dc.contributor.authorCorreia, I.J.
dc.contributor.authorMoreira, André F.
dc.date.accessioned2021-01-21T17:28:08Z
dc.date.available2021-01-21T17:28:08Z
dc.date.issued2020-12-29
dc.description.abstract3D tumor spheroids have arisen in the last years as potent tools for the in vitro screening of novel anticancer therapeutics. Nevertheless, to increase the reproducibility and predictability of the data originated from the spheroids it is still necessary to develop or optimize the techniques used for spheroids' physical and biomolecular characterization. Fluorescence microscopy, such as confocal laser scanning microscopy (CLSM), is a tool commonly used by researchers to characterize spheroids structure and the antitumoral effect of novel therapeutics. However, its application in spheroids' analysis is hindered by the limited light penetration in thick samples. For this purpose, optical clearing solutions have been explored to increase the spheroids' transparency by reducing the light scattering. In this study, the influence of agitation conditions (i.e., static, horizontal agitation, and rotatory agitation) on the ClearT and ClearT2 methods' clearing efficacy and tumor spheroids' imaging by CLSM was characterized. The obtained results demonstrate that the ClearT method results in the improved imaging of the spheroids interior, whereas the ClearT2 resulted in an increased propidium iodide mean fluorescence intensity as well as a higher signal depth in the Z-axis. Additionally, for both methods, the best clearing results were obtained for the spheroids treated under the rotatory agitation. In general, this work provides new insights on the ClearT and ClearT2 clearing methodologies and their utilization for improving the reproducibility of the data obtained through the CLSM, such as the analysis of the cell death in response to therapeutics administration.pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.doi10.3390/ijms22010266pt_PT
dc.identifier.urihttp://hdl.handle.net/10400.6/11052
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherMDPIpt_PT
dc.relationDevelopment of layer-by-layer polymeric microneedles for localized delivery of therapeutics to cervical cancer
dc.relation3D multicellular spheroids as high throughput platforms to screen novel combinatory therapies for treatment of pancreatic cancer
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/pt_PT
dc.subjectClearTpt_PT
dc.subjectClearT2pt_PT
dc.subjectConfocal microcopypt_PT
dc.subjectPropidium iodidept_PT
dc.subjectTumor spheroidspt_PT
dc.titleInfluence of ClearT and ClearT2 Agitation Conditions in the Fluorescence Imaging of 3D Spheroidspt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.awardTitleDevelopment of layer-by-layer polymeric microneedles for localized delivery of therapeutics to cervical cancer
oaire.awardTitle3D multicellular spheroids as high throughput platforms to screen novel combinatory therapies for treatment of pancreatic cancer
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/POR_CENTRO/SFRH%2FBD%2F109482%2F2015/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT//SFRH%2FBD%2F103507%2F2014/PT
oaire.citation.conferencePlaceSwisspt_PT
oaire.citation.issue1pt_PT
oaire.citation.startPage266pt_PT
oaire.citation.titleInternational Journal of Molecular Sciencespt_PT
oaire.citation.volume22pt_PT
oaire.fundingStreamPOR_CENTRO
person.familyNameda Rocha Costa
person.familyNameFélix Rodrigues
person.familyNameMiguel de Melo Diogo
person.familyNameJoaquim Sobreira Correia
person.familyNameMoreira
person.givenNameElisabete Cristina
person.givenNameAna Carolina
person.givenNameDuarte
person.givenNameIlídio
person.givenNameAndré Ferreira
person.identifierC2z5x04AAAAJ
person.identifierUMbJ1KMAAAAJ
person.identifier1161643
person.identifier.ciencia-idB314-4CF0-6633
person.identifier.ciencia-id3D14-BE84-E5CD
person.identifier.ciencia-id9C10-9BD8-634E
person.identifier.ciencia-idF610-7373-DC81
person.identifier.ciencia-id4C1E-012B-83CE
person.identifier.orcid0000-0002-0490-0095
person.identifier.orcid0000-0002-5493-8331
person.identifier.orcid0000-0001-9984-3603
person.identifier.orcid0000-0003-1613-9675
person.identifier.orcid0000-0002-0604-2506
person.identifier.scopus-author-id55805850100
person.identifier.scopus-author-id57202076016
person.identifier.scopus-author-id56266511300
person.identifier.scopus-author-id7003557499
person.identifier.scopus-author-id56401521300
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT
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