Flow Cytometric Analysis of GFP Expression in Mammalian Cells

Abstract

In the jellyfish Aequorea victoria, light is produced when energy is transferred from the Ca²⁺-activated photo protein aequorin to green fluorescent protein or GFP (Fig. 1).^1-3 This process occurs in specialized photogenic cells located at the base of jellyfish umbrella, where each protein is found at very high concentrations. The cloning of the wild-type GFP gene (wt GFP)^4,5 and its subsequent expression in heterologous systems^6,7 has established GFP an important reporter protein for the analysis of gene expression and protein localization in a wide variety of experimental designs. When expressed in either eukaryotic or prokaryotic cells and illuminated by blue or UV light, wt GFP emits a bright green fluorescent signal which is easily detected by fluorescence microscopy, flow cytometry, or other fluorescence imaging techniques. GFP fluorescence is species-independent and does not require additional cofactors, substrates, or gene products — the protein is “naturally fluorescent”. Moreover, detection of GFP and its variants can be performed in living samples, and is ideally suited to real time analysis of molecular events.