Approaches to Optimizing High Content Confocal Microscopy

Sammanfattning: 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.