I\'d like to be able to measure the activity of ? -galactosidase in living cells

Question

I'd like to be able to measure the activity of ?-galactosidase in living cells with simple optical (maybe fluorescence) microscopy. Ideally I'd like to do a minimum of genetic engineering, and use this assay with strains I already have (that have a WT lactose system), i.e. a fluorescent lactose-mimic would be ideal. Additionally, it would be nice if the lifetime of the fluorescent byproduct of ?-gal activity was short, so I could detect a decrease in ?-gal activity as well as an increase. Does such a fluorescent analog exist? I'm hoping to look at the switch from lactose to glucose utilization under the microscope, so I want lactose-using cells to light up and glucose-using cells to be dark, or vice versa. It does not have to be fully quantitative--a qualitative sense is fine.

Explanation / Answer

I found this paper.

Zhang GJ et al. (2009) In vivo optical imaging of LacZ expression using lacZ transgenic mice. Assay Drug Dev Technol.7:391-9. doi: 10.1089/adt.2009.0195.

Abstract

beta-Galactosidase (beta-gal) (encoded by the lacZ gene) has been widely used as a transgene reporter enzyme. The ability to image lacZ expression in living transgenic animals would further extend the use of this reporter. It has been reported that 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one)-beta-d-galactopyranoside (DDAOG), a conjugate of beta-galactose and 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one), is not only a chromogenic lacZ substrate but that the cleavage product has far-red fluorescence properties detectable by in vivo imaging. In an attempt to noninvasively image lacZ expression in vivo, we applied fluorescence imaging to a G protein-coupled receptor (GPR56), knockout (KO) mouse model, in which the lacZ gene is introduced in the GPR56 locus. Compared to wild-type (WT) mice, GPR56KO/LacZ mice showed three- to fourfold higher fluorescence intensity in the head area 5 min after tail-vein injection of DDAOG. beta-Gal staining in sections of whole brain showed strong lacZ expression in homozygotes, but not in WT mice, consistent with lacZ activity detected by in vivo imaging. The kidneys were also visualized with fluorescence imaging both in vivo and ex vivo, consistent with beta-gal staining findings. Our results demonstrate that fluorescence imaging can be used for in vivo real-time detection of lacZ activity by fluorescence imaging in lacZ transgenic mice. Thus, this technology can potentially be used to noninvasively image changes of certain endogenous molecules and/or molecular pathways in transgenic animals.

The work in the paper is cited in a more recent paper which reports beta-gal-based chemiluminescent imaging in mice.

It's unclear if any of this would be successful at a single bacterial cell level.

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