Simultaneous detection of DsRed2-tagged and EGFP-tagged human beta-interferons in the same single cells.
The red fluorescent protein DsRed2 is a useful fusion tag for various proteins, together with the enhanced green fluorescent protein (EGFP). These chromoproteins have spectral properties that allow simultaneous distinctive detection of tagged proteins in the same single cells by dual color imaging. We used them for tagging a secretory protein, human interferon-beta (IFN-beta). Expression plasmids for human IFN-beta tagged with DsRed2 or with EGFP at the carboxyl terminal were constructed and their coexpression was examined in Mardin-Darby canine kidney epithelial cells. Although maturation of DsRed2 for coloration was slow and the color intensity was weak compared with EGFP, low temperature treatment (20 degrees C) allowed DsRed2-tagged human IFN-beta to be detected in the cells using color imaging. Consequently, the two chimeric proteins were shown to be colocalized in the same single cells by dual color confocal microscopy. This approach will be useful for investigating subcellular localization of not only cell resident proteins but also secretory proteins.
Maruyama
Masato
M
Department of Biopharmaceutics and Drug Metabolism, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Nishio
Teruko
T
Yoshida
Toyokazu
T
Ishida
Chisaki
C
Ishida
Kayo
K
Watanabe
Yoshihiko
Y
Nishikawa
Makiya
M
Takakura
Yoshinobu
Y
D000818:Animals; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004285:Dogs; D004847:Epithelial Cells / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006801:Humans; D016899:Interferon-beta / Q000378:metabolism*; D008164:Luminescent Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D000097573:Red Fluorescent Protein
D049452:Green Fluorescent Proteins; D016899:Interferon-beta; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D000097573:Red Fluorescent Protein