title: Immunofluorescence and fluorescent-protein tagging show high correlation for protein localization in mammalian cells.
abstract: Imaging techniques such as immunofluorescence (IF) and the expression of fluorescent protein (FP) fusions are widely used to investigate the subcellular distribution of proteins. Here we report a systematic analysis of >500 human proteins comparing the localizations obtained in live versus fixed cells using FPs and IF, respectively. We identify systematic discrepancies between IF and FPs as well as between FP tagging at the N and C termini. The analysis shows that for 80% of the proteins, IF and FPs yield the same subcellular distribution, and the locations of 250 previously unlocalized proteins were determined by the overlap between the two methods. Approximately 60% of proteins localize to multiple organelles for both methods, indicating a complex subcellular protein organization. These results show that both IF and FP tagging are reliable techniques and demonstrate the usefulness of an integrative approach for a complete investigation of the subcellular human proteome.
authors: KTH Royal Institute of Technology, Science for Life Laboratory, School of Biotechnology, Stockholm, Sweden.
authors: Charlotte
authors: C
authors: Stadler
authors: Elton
authors: E
authors: Rexhepaj
authors: Vasanth R
authors: VR
authors: Singan
authors: Robert F
authors: RF
authors: Murphy
authors: Rainer
authors: R
authors: Pepperkok
authors: Mathias
authors: M
authors: Uhlén
authors: Jeremy C
authors: JC
authors: Simpson
authors: Emma
authors: E
authors: Lundberg
mesh_terms: D000818:Animals; D002522:Chlorocebus aethiops; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D025941:Protein Interaction Mapping; D021381:Protein Transport / Q000502:physiology*; D013194:Staining and Labeling / Q000379:methods*; D014709:Vero Cells
chemical_list: D008164:Luminescent Proteins