SENSITIVE AND RAPID DETECTION OF BETA-GALACTOSIDASE EXPRESSION IN INTACT-CELLS BY MICROINJECTION OF FLUORESCENT SUBSTRATE

Bacterial beta-galactosidase, coded by lacZ, is a widely used reporter for studies of transcriptional activity of eukaryotic promoters at the single cell level. Unfortunately, current detection methods, Like X-gal cytochemistry, are slow, have suboptimal sensitivity, and are incompatible with cell survival. By a novel approach based on microinjection into cells of the fluorogenic substrate 5-chloromethylfluorescein di-beta-D-galactopyranoside lacZ gene expression was detected without affecting cell viability or proliferative capacity. The method was far more sensitive than the conventional X-gal cytochemistry in all cell systems tested (primary hepatocytes, fibroblasts, and glioma cells). Results were obtained within seconds to minutes after injection, and cells remained fluorescent for hours. (C) 1995 Academic Press, Inc.