Use of fluorescein-di-beta-D-galactopyranoside (FDG) and C12-FDG as substrates for beta-galactosidase detection by flow cytometry in animal, bacterial, and yeast cells.
Fluorescein-di-beta-D-galactopyranoside (FDG) was found to be a useful substrate for beta-galactosidase detection by flow cytometry in gram-negative bacteria, since it entered viable cells and gave a fluorescence emission proportional to the enzymatic activity. C12-FDG, a more lipophilic derivative, gave a very poor signal because of the lack of penetration. On the contrary, C12-FDG was more sensitive than FDG for beta-galactosidase activity determinations in animal cells. In contrast to previous reports, C12-FDG did not enter viable yeast cells, so that the use of the substrate required cell permeabilization. Without this treatment, C12-FDG penetrates only nonviable yeast cells that may occur in populations expressing beta-galactosidase.
Plovins
A
A
Departamento de Microbiología II, Facultad de Farmacia, Universidad Complutense, Madrid, Spain.
Alvarez
A M
AM
Ibañez
M
M
Molina
M
M
Nombela
C
C
D000818:Animals; D002460:Cell Line; D002463:Cell Membrane Permeability; D002469:Cell Separation; D004926:Escherichia coli / Q000201:enzymology; D005434:Flow Cytometry / Q000379:methods*; D005452:Fluoresceins / Q000378:metabolism*; D005697:Galactosides / Q000378:metabolism*; D006801:Humans; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012680:Sensitivity and Specificity; D001616:beta-Galactosidase / Q000032:analysis*
D005452:Fluoresceins; D005697:Galactosides; C038848:fluorescein-digalactoside; D001616:beta-Galactosidase