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Approaches to Optimizing High Content Confocal Microscopy
KTH, School of Engineering Sciences (SCI), Applied Physics. (Cellens fysik)
2009 (English)Licentiate thesis, monograph (Other academic)
Abstract [en]

This thesis presents two methods for improving high contentconfocal microscopy.

The author's part in the first, "Toward a confocal subcellular atlasof the human proteome" was automating image capture of foursimultaneously tagged structures in cells in 96 well plates. In total,thousands of images of hundreds of proteins in cells. The authorwas also part of deciding which imaging methods should be used tomaximize image information content and quality, given the limitedtime available per well in order to scan large numbers of wells.

The second project, "Improved water permeability measurementsbased on fluorescence normalization" involves increasing the sensitivityof measurements of protein function by normalizing with anestimate of the amount of protein in the cell - the fluorescentsignal of a co-transfected protein. This could lead to achievingsufficient confidence in measurements with fewer experiments(thus increasing the information content in each experiment). Asurprisingly high level of noise in the relationship between thefluorescent signal and the protein function was measured.

Abstract [sv]

Denna avhandling presenterar två projekt för att förbättrametoder för experiment med stora informationsmängderbaserade på konfokalmikroskopi.

Författarens del i det första projektet, "Toward a ConfocalSubcellular Atlas of the Human Proteome" (Mot en konfokal,subcellulär atlas av det mänskliga proteomet) var att automatiserabildinsamlingen av fyra samtidigt inmärkta strukturer i celler iplattor med 96 brunnar. Sammanlagt togs tusentals bilder avhundratals proteiner i celler. Författaren var även del i att fastställavilka bildinsamlingsmetoder som skulle användas för att maximeramängd och kvalitet på bild-informationen givet den begränsade tidper brunn som var tillgänglig för att kunna avbilda många brunnar.

Den andra studien, "Improved water permeability measurementsbased on fluorescence normalization" (Förbättrade vattenpermeabilitetsmätningargenom normalisering av fluorescens) syftade till att ökakänsligheten hos mätningar av proteiners funktion genom attnormalisera mätningarna med signalen från fluorescensen från ettkotransfekterat protein. Det skulle kunna leda till att nå tillräckligtillförlitlighet i mätresultaten med färre experiment (därmed ökainformationsinnehållet i varje experiment). En förvånansvärt högbrusnivå i förhållandet mellan fluorescenssignalen ochproteinfunktionen uppmättes

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2009. , 50 p.
Series
TRITA-FYS, ISSN 0280-316X ; 2009:11
National Category
Physical Sciences
Identifiers
URN: urn:nbn:se:kth:diva-10691ISBN: 978-91-7415-280-7 (print)OAI: oai:DiVA.org:kth-10691DiVA: diva2:225234
Presentation
2009-06-08, Sal FA32, AlbaNova, Roslagstullsbacken 21, Stockholm, 13:00 (English)
Opponent
Supervisors
Available from: 2009-06-25 Created: 2009-06-25 Last updated: 2017-05-23Bibliographically approved

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