Objective: A recently developed assay to study drug-modulating effects on gap-junction permeability is analysed. The assay is based on electroporation of a large number of cells grown into a confluent monolayer. The subsequent intercellular spread of loaded dye to non- electroporated parts of the monolayer estimates relative intercellular coupling. It is not clear however, how the measures used so far of relative changes in fluorescence intensity, length constants and manual cell counting relates to the underlying change in physical permeability, as expressed by the permeability coefficient. Methods: We present a mathematical model of a large monolayer of coupled cells. This simulated setup has properties similar to the electroporation assay. Results: The mathematical model enables estimation of the permeability coefficient. The simulations show that a major problem with the above-mentioned measures is that they do not display proportionality with time, i.e. over a broad range of time they seem unable to converge towards a single value for the underlying permeability coefficient. We argue that the most commonly used measure - manual cell counting – is inferior due to inter-observer differences and the risk of bias. In addition, new measures deduced from Fick's law of diffusion are presented which directly relate the observed spread of dye to the permeability coefficient. Conclusion: In terms of the underlying permeability coefficient, the hitherto used measures give a relatively poor degree of quantification. In contrast, the presented new measures provide direct information on the inter- cellular permeability coefficient. Hence, these new measures may provide additional and more precise information as to the permeability modulating effect of various substances.