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PubMed · 10752489

Green fluorescent protein as a quantitative tool.

N J Hack·B Billups·P B Guthrie·J H Rogers·E M Muir·T N Parks·S B Kater
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Abstract

Manipulating the expression of a protein can provide a powerful tool for understanding its function, provided that the protein is expressed at physiologically-significant concentrations. We have developed a simple method to measure (1) the concentration of an overexpressed protein in single cells and (2) the covariation of particular physiological properties with a protein's expression. As an example of how this method can be used, teratocarcinoma cells were transfected with the neuron-specific calcium binding protein calretinin (CR) tagged with green fluorescent protein (GFP). By measuring GFP fluorescence in microcapillaries, we created a standard curve for GFP fluorescence that permitted quantification of CR concentrations in individual cells. Fura-2 measurements in the same cells showed a strong positive correlation between CR-GFP fusion protein expression levels and calcium clearance capacity. This method should allow reliable quantitative analysis of GFP fusion protein expression.

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AnimalsCalbindin 2CalciumFluorescent Antibody TechniqueGreen Fluorescent ProteinsHumansIndicators and ReagentsLuminescent ProteinsMaleS100 Calcium Binding Protein GTeratocarcinomaTesticular NeoplasmsTumor Cells, CulturedJournal ArticleResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, P.H.S.

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N J Hack, B Billups, P B Guthrie, J H Rogers, E M Muir, T N Parks, S B Kater. 2000-02-15. Green fluorescent protein as a quantitative tool.. https://doi.org/10.1016/s0165-0270(99)00178-8

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