What is lacZ staining?
LacZ is a frequently used reporter gene, encoding for the protein beta-galactosidase in cultured cells, which appear blue when the cultured cells are grown on a medium containing X-gal analog. The staining solution is made of X-gal (200 mg/ml), MgCl2 (1M), K ferri-cyanide (50mM), K ferro-cyanide (50mM), and PBS.
How do you detect lacZ?
The expression of lacZ can be detected by enzyme-based histochemical staining using chromogenic substrates such as 5-bromo-4-chloro-3-indolyl-β-d-galactoside (X-gal).
What does beta gal Stain for?
The β-Gal Staining Kit allows you to determine the percentage of transfected cells expressing lacZ. β-galactosidase catalyzes the hydrolysis of X-gal, producing a blue color that can be easily visualized under a microscope. …
What is beta-galactosidase assay?
Additional Information. The β-Gal Assay Kit provides the reagents required to quickly measure the levels of active β-galactosidase expressed in cells transfected with plasmids expressing the lacZ gene. lacZ is a bacterial gene often used as a reporter construct in eukaryotic transfection experiments.
Is lacZ a fluorescent?
Unlike the above two types of markers, LacZ does not emit light. Although highly sensitive, LacZ/X-gal staining is not quantitative, and does not provide the high resolution possible with fluorescent or bioluminescent proteins.
Why is lacZ important?
It played a central role in Jacob and Monod’s1 development of the operon model for the regulation of gene expression. Also, its ability to signal its presence by producing an easily recognizable blue reaction product has made it a workhorse in cloning and other such molecular biology procedures.
How to prepare lacZ histochemistry solution for staining?
If lacZ histochemistry is to be followed by antibody staining, a 0.4% paraformaldehyde fixation solution can be used instead. 3. Rinse the fixed tissues in detergent rinse three times for 15-30 minutes at room temperature. 4. Incubate the samples in embryo staining solution.
How do I stain for β-galactosidase (lacZ) activity in mouse embryos?
Whole mouse embryos are stained for β-galactosidase ( lacZ) activity using X-gal. A protocol for Staining Frozen Mouse Embryo Sections for β-Galactosidase (lacZ) Activity is also available. If staining is to be performed for more than 1 hour, add Tris-Cl (pH 7.3) to the staining solution to a final concentration of 20 mM.
How long does it take to stain cells with lacZ?
Stain cells (embryos) 4 hours to overnight at 37*C depending on level of LacZ activity. 5. After staining, pour off stain, replace with wash buffer.
Do I need to counterstain lacZ staining?
When sections are viewed by dark-field microscopy, the lacZ stain is pink and the contrast with surrounding tissues is good; therefore, no counterstaining is necessary. doi:10.1101/pdb.prot4725 Cold Spring Harb Protoc 2007.