LacZ beta Galactosidase Intracellular Detection Kit (ab189816)
Key features and details
- Platform: Microplate reader, Flow cytometer
Overview
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Product name
LacZ beta Galactosidase Intracellular Detection Kit
See all beta galactosidase kits -
Product overview
Abcam’s LacZ beta Galactosidase Intracellular Detection Kit (ab189816) uses the β -galactoside analog fluorescein di-ß-D-galactopyranoside (FDG) in a protocol that sensitively distinguishes lacZ+ cells from lacZ- cells. Although chromogenic assays of ß-galactosidase activity (i.e. X-Gal) are useful, the recent application of the fluorogenic substrate fluorescein di-ß-D-galactopyranoside (FDG) has been shown to be several orders of magnitude more sensitive. Because of this high sensitivity, use of FDG allows quantitation of lacZ expression in single, viable eukaryotic cells, whereas other assays often result in dead cells. In addition, because of its high water solubility and detection limits, the FDG substrate has found extensive use in automated ELISA type assay systems.
Emission: 512 nm.
Excitation: 488 nm.
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Notes
One of the most common reporter genes used in molecular biology applications is the E. coli lacZ gene that codes for an active subunit of β-galactosidase in vivo. Since this enzyme is generally absent in normal mammalian, yeast, some bacterial and even plant cells, it can be detected at very low levels, and since the enzyme has a wide substrate specificity, monitoring lacZ expression (and therefore co-expressed genes or promoter efficiency) has become routine to the point of detection of as few as 5 copies of β-galactosidase per cell.
Abcam’s LacZ beta Galactosidase Intracellular Detection Kit (ab189816) provides reagents and protocol to perform up to 250 assays.
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Platform
Microplate reader, Flow cytometer
Properties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components 250 tests Chloroquine 1 x 1ml Fluorescent Substrate Reagent 1 x 1ml PETG 1 x 1ml Reference Standard 1 x 1ml -
Research areas
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Relevance
Beta galactosidase is a hydrolase enzyme that cleaves beta-linked terminal galactosyl residues from gangliosides, glycoproteins, and glycosaminoglycans. Beta galactosidase is an essential enzyme in the human body. Deficiencies in the protein can result in galactosialidosis or Morquio B syndrome. Senescent cells display senescence-associated expression of beta galactosidase activity. -
Cellular localization
Isoform 1: Lysosome. Isoform 2: Cytoplasm, perinuclear region. Note=Localized to the perinuclear area of the cytoplasm but not to lysosomes. -
Alternative names
- Acid beta galactosidase
- Beta galactosidase 1
- EBP
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Images
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Example data
Two adherent mouse fibroblast tumor cell lines, CRE BAG 2 (lacZ stable transformants), and NIH 3T3 (lacZ negative) were cultured to 50% confluence in 12-well tissue culture plates (clear, flat bottom). Media was replaced with Dulbecco’s Modified Eagles Medium (DMEM) containing no serum or antibiotics (1 mL). Plate wells were also prepared containing medium only (no cells). Cells were incubated (37°C, 5% CO2) for 24 hours. Fluorescent Substrate Reagent was added to all wells to a final concentration of 500 μM (50 μL). Fluorescence was recorded using 485nm excitation and 535nm emission filters. Readings were taken at 1, 3, and 5 hour intervals. Assays were performed in triplicate and averaged, error bars represent standard error.
