Open AccessA novel method using confocal laser scanning microscopy for sensitive measurement of P‐glycoprotein‐mediated transport activity in Caco‐2 cells
Author(s) -
Wakuda Hirokazu,
Nejime Namie,
Tada Yukari,
Kagota Satomi,
Fahmi Odette A.,
Umegaki Keizo,
Yamada Shizuo,
Shinozuka Kazumasa
Publication year - 2011
Publication title -
journal of pharmacy and pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 118
eISSN - 2042-7158
pISSN - 0022-3573
DOI - 10.1111/j.2042-7158.2011.01294.x
Subject(s) - confocal , rhodamine 123 , confocal microscopy , fluorescence microscope , microscopy , fluorescence , rhodamine , laser scanning , laser microscopy , microscope , confocal laser scanning microscope , chemistry , biophysics , laser , analytical chemistry (journal) , materials science , chromatography , biology , biomedical engineering , biochemistry , optics , medicine , pathology , microbiology and biotechnology , physics , multiple drug resistance , antibiotics
Objectives The aim of this study was to use time‐lapse confocal laser scanning microscopy to establish a more sensitive and specific method for evaluating P‐glycoprotein activity in Caco‐2 cells. Methods The change in the fluorescence of residual rhodamine 123 at the apical and central regions of Caco‐2 cells was measured in the presence of digoxin or St John's wort by using time‐lapse confocal laser scanning microscopy. The data were compared with measurements made using conventional techniques, a fluorescence microplate reader and a fluorescence microscope. Key findings The percentage decrease of rhodamine 123 caused by 10 µ m digoxin or 0.1 µg/ml St John's wort was significantly larger in the apical region of the Caco‐2 cell than in the central region or in the whole cell. The digoxin‐induced inhibition in the apical region as measured by time‐lapse confocal laser scanning microscopy was greater than that measured in the whole cell by a microplate reader or a fluorescence microscope. Conclusions The assay of residual rhodamine 123 in the apical region of Caco‐2 cells by confocal laser scanning microscopy was more sensitive than the conventional methods using a microplate reader or fluorescence microscopy. It will be a valuable screening tool for studying both the inhibition and induction of P‐glycoprotein activity.