European Biophysics Journal

, Volume 41, Issue 3, pp 341–352 | Cite as

The statistics of protein expression ratios for cellular fluorescence studies

  • Elizabeth M. Smith
  • Joachim D. Mueller
Original Paper


Fluorescence studies of cellular protein-protein interactions commonly employ transient cotransfection to express two proteins carrying distinct fluorescent labels. Because transiently transfected cells differ significantly in their expression level, the concentration ratio of the two expressed proteins varies, which in turn influences the measured fluorescence signal. Knowledge of the statistics of protein expression ratios is of considerable interest both from a fundamental point of view and for cellular fluorescence studies. Despite the perceived randomness of transient transfection, we were able to develop a quantitative model that describes the average and distribution of the protein expression ratio from a cell population. We show that the expression ratio is proportional to the molar plasmid ratio and relate the distribution to the finite number of active plasmids in the cell. The process of cationic lipid-mediated transfection is explored in more detail. Specifically, the influence of lipoplexes on the statistics of the expression ratio is examined. We further demonstrate that the transfection model provides a quantitative description of fluorescence fluctuation experiments, where only a fraction of the proteins are labeled.


Brightness analysis Lipoplexes Fluorescence fluctuation spectroscopy Fluorescent proteins 



This work was supported by grants from the National Institutes of Health (GM64589) and the National Science Foundation (PHY 0346782). We thank Dr. Joseph Albanesi for kindly providing the Endophilin A2 plasmid.


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Copyright information

© European Biophysical Societies' Association 2012

Authors and Affiliations

  1. 1.School of Physics and AstronomyUniversity of MinnesotaMinneapolisUSA
  2. 2.Institute for Molecular VirologyUniversity of MinnesotaMinneapolisUSA
  3. 3.Department of Biomedical EngineeringUniversity of MinnesotaMinneapolisUSA

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