Updated: September 25, 2019

For research use only

lycoGREEN™-βGal
Table 1. Product information
Catalog no.

Product nameAmountStorage upon receipt

Stability

GC611

GlycoGREEN™-βGal

30 nmol×5≤−20°C, keep desiccated1 year (when unopenedand protected from light.

and stored as described.)

Ă-̀

1.

GlycoGREEN ™ -βGal is a fluorescent probe that can detect β-galactosidase (βGal) activity by green
fluorescence. This probe is suitable for live cell imaging, screening using a microplate reader, flow cytometry
analyses, and detection of βGal activity in fixed cells. It can be also used for the detection of increase in theβGal activity in senescent cells or in cancer cells.

Figure 1. (left) Absorbance and fluorescence spectra of GlycoGREEN™-βGal.
(right) Fluorescent spectra before and after the reaction with βGal. Fluorescenceincreases >200-fold upon reaction with βGal.

■ Storage of the reagent
The reagent is shipped as a dried powder in a nitrogen gas filled tube. Upon receipt, store at ≤−20°C, keep
desiccated and protected from light. Dissolve the reagents to DMSO just before the use. We provide nowarranty for the reagents which was stored as a solution.

■ Preparation of the reagent
1. GlycoGREEN™-βGal is a colorless powder. Centrifuge the tube before opening the cap because a part ofthe powder might be adhered on the cap.
2. Warm the vial to the room temperature before opening the cap. Add 30 μl of DMSO to one vial to prepare
1 mM stock solution. Dissolve the powder completely by careful pipetting. Solution should also be acolorless liquid.

2.
■ Cell imaging example 1—Detection of LacZ gene expression in HEK293 cells① Prepare HEK293 cells with and without expressing LacZ gene (LacZ+/-) on glass bottom dishes. Culturecells in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 8% FBS, penicillin andstreptomycin at 37oC, 5% CO2 for overnight.
② Dilute GlycoGREEN™-βGal with the culture medium to be 1 μM (cell staining solution).
③ Remove the culture medium in the glass bottom dishes, add the cell staining solution, and incubate at 37oC
for 15 minutes.
④ Remove the cell staining solution, rinse the cells with HBSS. Observe cells in HBSS or in other observationbuffer/medium without phenol red, which may increase fluorescence backgrounds.
※ Instead of the above step 4, cells can be observed after fixation. For fixation, remove the medium, rinse
the cells with culture medium, and fix cells with 3% paraformaldehyde in PBS for 15 minutes.
※ Optimal concentration of GlycoGREEN™-βGal and reaction time may vary on cell types and conditions.
■ Cell imaging example 2—Staining after cell fixation① Prepare HEK293 LacZ+/- cells in glass bottom dishes and culture in DMEM + 8% FBS at 37oC, 5% CO2for overnight.
② Remove the medium, rinse the cells with culture medium, and fix cells with 3% paraformaldehyde in PBS
for 15 minutes.
③ Remove the staining solution and rinse cells with PBS for 3 times.
④ Dilute GlycoGREEN™-βGal with HBSS to prepare 1 μM solution (cell staining solution). Replace thebuffer to this solution and incubate at 37oC for 15 minutes.
⑤ Remove the culture medium and rinse cells with PBS or HBSS 2 times. Observe cells with fluorescencemicroscopy.
※ Longer fixation or higher concentration of aldehyde decrease the enzyme activity and reactivity of thereagent.
■ Cell imaging example 3—Distinguishing cancer cell lines and non-tumor cells① Culture non-tumor cells (ex. HUVEC cells) and a cancer cell line (ex. OVCAR5, HeLa, HepG2) in glassbottom dishes to reach 50-70% confluent.
② Dilute GlycoGREEN™-βGal with the culture medium to be 1 μM (cell staining solution).
③ Remove the medium from each dish, add cell staining solution, and incubate at 37oC for 2 hours.
④ Remove the cell staining solution, rinse cells with HBSS. Observe cells by fluorescence microscopy.
※β-- -■ Flow cytometric analysis① Prepare HEK293 LacZ+/- cells in glass bottom dishes and culture in DMEM + 8% FBS at 37oC, 5% CO2for overnight② Dilute GlycoGREEN™-βGal with the culture medium to 1 μM (cell staining solution).
③ Remove the culture medium in the glass bottom dishes, add the cell staining solution, and incubate at37oC for 15 minutes.

④ Remove the cell staining solution and rinse the cells with PBS. Remove PBS and add Trypsin/EDTA toeach dish and incubate until the cells have become detached.
⑤ Transfer cell suspension to a sterile centrifuge tube and centrifuge at 600×g, 5 minutes to collect cells.
Remove the media and resuspend to PBS.
⑥ Pass cells through a cell strainer to eliminate clumps and debris. Analyze by flow cytometry by following
the instruction of the system.
※β-■ Fluorescent observation
For excitation, 488 nm laser such as argon laser is appropriate. In fluorescence microscope, blue excitation
filter cube such as B-2A, FITC (NIKON), U-FBW, U-FBWA (OLYMPUS), or filters for FITC/GFP can be used.
For flow cytometry, filter set for FITC is appropriate.

■ References

Table 2. Related Products
Catalog no.

Product nameMajor applications™-βDetection of β-galactosidase activity by yellow fluorescence. For live cell
imaging and high content screening.

-®

-β

For screening of β-galactosidase activity by microplate reader.

-®

-β

For screening of β-glucosidase activity by microplate reader.

-®

-β

For screening of β-glucuronidase activity by microplate reader.

EAREE Bldg. 5F, Kita 8 Nishi 18-35-100, Chuo-ku, Sapporo 060-0008, JAPANTEL: +81-11-624-5860
https://goryochemical.com/