Characterization of the activity of β-galactosidase from and in fixed and non-fixed tissues.

β-Galactosidase encoded by the gene, is widely used as a reporter molecule in molecular biology in a wide variety of animals. β-Galactosidase retains its enzymatic activity in cells or tissues even after fixation and can degrade X-Gal, a frequently used colormetric substrate, producing a blue color. Therefore, it can be used for the activity staining of fixed tissues. However, the enzymatic activity of the β-galactosidase that is ectopically expressed in the non-fixed tissues of animals has not been extensively studied. Here, we report the characterization of β-galactosidase activity in tissues with and without fixation in various experimental conditions comparing the activity of two evolutionarily orthologous β-galactosidases derived from the . and genes. We performed quantitative analysis of the activity staining of larval imaginal discs and an assay using larval lysates. Our data showed that both . and β-galactosidase can be used for cell-type-specific activity staining, but they have their own preferences in regard to conditions. . β-galactosidase showed a preference for neutral pH but not for acidic pH compared with β-galactosidase. Our data suggested that both . and β-galactosidase show enzymatic activity in the physiological conditions of living animals when they are ectopically expressed in a desired specific spatial and temporal pattern. This may enable their future application to studies of chemical biology using model animals.