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SPiDER-ßGal

Item # Unit Size
SG02-10
20 µg × 3

For Research Use Only Products



Storage Condition : 0-5oC
Shipping Condition : ambient temperature
CAS# : 1824699-57-1

Product Description
The gene of β-galactosidase from E. coli is widely used as a reporter gene assay marker. Although X-gal is well known reagent to detect β-galactosidase in cell or tissue samples, the assay using these reagents require to fix cells or tissues due to the poor cell-permeability. In addition, so far developed the assay using fluorescence reagents cannot clearly differentiate β-galactosidase-expressed cells or regions.
To overcome these issues, Urano, Kamiya and co-workers have successfully developed SPiDER-βGal. SPiDER-βGal ideally possesses cell-permeability and the ability to retain in intracellular region.1)
By the enzymatic reaction, SPiDER-βGal immediately forms a quinone methide that acts as electrophile when proteins containing nucleophilic functional groups nearby the molecules. By the probe undergoes the reaction with a protein, the conjugates become fluorescent compounds. Thus, SPiDER-βGal allows a single-cell analysis because it does self-immobilizing to the intracellular proteins.


Usage Example
Fluorescence microscopic detection of β-galactosidase-expressed cells

β-galactosidase-expressed cells (HEK/LacZ cells) were clearly observed in fluorescence imaging. In addition, the result was not changed by fixing the cells.

  1. HEK cells at 5 × 105 cells/ml (500 μl) and HEK/LacZ cells at 5 × 105 cells/ml (500 μl) were seeded in a 35 mm dish in DMEM (10% fetal bovine serum, 1% penicillin-streptmycin) and cultured overnight in a 5% CO2 incubator at 37oC.
  2. The cells were washed with 2 ml of Hanks’ HEPES buffer twice.
  3. SPiDER-βGal working solution (2 ml) was added to the culture dish. The cells were then incubated for 15 minutes at 37oC.
  4. After the supernatant was removed, the cells were washed Hanks’ HEPES buffer (2 ml) twice.
  5. Hanks’ HEPES buffer (2 ml) were added and the cells observed under a fluorescence microscope. (Fig. 3A)
  6. After the supernatant was removed, 4% paraformaldehyde (PFA) /PBS solution (2 ml) was added to the culture dish. The cells were then incubated for 15 minutes at room temperature.
  7. After 4% PFA/PBS solution was removed, the cells were washed Hanks’ HEPES buffer (2 ml) twice.
  8. Hanks’ HEPES buffer (2 ml) were added and the cells observed under a fluorescence microscope. (Fig. 3B)

Flow cytometric detection of β-galactosidase-expressed cells
  1. HEK cells at 5 × 105 cells/ml (500 μl) and HEK/LacZ cells at 5 × 105 cells/ml (500 μl) were mixed in a microtube.
  2. SPiDER-βGal DMSO stock solution (1 μl) was added to the tube. The cells were then incubated 15 minutes at 37oC .
  3. The cells were analyzed under a flow cytometer. (488 nm excitation, 530/30 nm bandpass filter)


β-galactosidase-expressed cells (HEK/LacZ cells) were clearly differentiate from HEK cells in flow cytometry data analysis.