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Xite™ Green beta-D-galactopyranoside

Xite™ Green beta-D-galactopyranoside is a fluorogenic substrate for beta-galactosidase (β-gal). Compared to the existing beta-galactosidase substrates (e.g., the commonly used FDG), it has much better cell permeability. Xite™ Green beta-D-galactopyranoside readily enters cells where it gets cleaved by β-gal, producing Xite™ Green, a strongly fluorescent product. The strongly fluorescent Xite™ Green is well retained in cells, making it easy to be detected with a flow cytometer and fluorescence microscope. Xite™ Green beta-D-galactopyranoside provides a simple and sensitive tool to detect beta-galactosidase activity. Xite™ Green beta-D-galactopyranoside might be used as a simple tool for measuring cellular senescence in cells since β-gal has been identified as a reliable marker for cellular senescence.

Example protocol

AT A GLANCE

Protocol summary
  1. Treat samples as desired
  2. Prepare and add Xite™ Green beta-D-galactopyranoside working solution to samples
  3. Incubate samples at 37 °C for 15 to 45 minutes
  4. Monitor the fluorescence intensity using flow cytometer with 530/30 nm filter (FITC channel) or using fluorescence microscopy with FITC filter set 

PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

Xite™ Green beta-D-galactopyranoside stock solution
Add appropriate amount of DMSO into Xite™ Green beta-D-galactopyranoside to make 2-5 mM Xite™ Green beta-D-galactopyranoside stock solution. Note: Store the unused Xite™ Green beta-D-galactopyranoside stock solution at -20 °C in single use aliquots.

PREPARATION OF WORKING SOLUTION

Xite™ Green beta-D-galactopyranoside working solution
Prepare 1-20 µM of Xite™ Green beta-D-galactopyranoside working solution in buffer of your choice. Note: Xite™ Green beta-D-galactopyranoside working solution should be used promptly. Note: The concentration of the Xite™ Green beta-D-galactopyranoside should be optimized for different cell types and conditions.

SAMPLE EXPERIMENTAL PROTOCOL

The following protocol can be used as a guideline and should be optimized according to the needs.
  1. Treat your samples as desired.
  2. Remove the treatment and wash the cells with buffer of your choice such as DPBS. Note: For selectively tracking β-Gal in live cells, cells can be treated with Bafilomycin A1 for blocking endogenous β-Gal. Optimum concentration of Bafilomycin A1 may vary on type of cells.
  3. Add Xite™ Green beta-D-galactopyranoside working solution for 15-45 minutes and incubate the samples at 37 °C incubator. Note: Optimal time for incubation needs to be determined experimentally.
  4. Remove the working solution and wash cells with buffer of your choice.
  5. Resuspend the cells in buffer of your choice and monitor the fluorescence intensity with flow cytometer using 530/30 nm filter (FITC channel) or fluorescence microscope with FITC filter set. 

References

View all 18 references: Citation Explorer
Novel fluorescent probe for rapid and ratiometric detection of β-galactosidase and live cell imaging.
Authors: Chen, Xiangzhu and Zhang, Xueyan and Ma, Xiaodong and Zhang, Yuanyuan and Gao, Gui and Liu, Jingjing and Hou, Shicong
Journal: Talanta (2019): 308-313
Targeting senescence improves angiogenic potential of adipose-derived mesenchymal stem cells in patients with preeclampsia.
Authors: Suvakov, Sonja and Cubro, Hajrunisa and White, Wendy M and Butler Tobah, Yvonne S and Weissgerber, Tracey L and Jordan, Kyra L and Zhu, Xiang Y and Woollard, John R and Chebib, Fouad T and Milic, Natasa M and Grande, Joseph P and Xu, Ming and Tchkonia, Tamara and Kirkland, James L and Lerman, Lilach O and Garovic, Vesna D
Journal: Biology of sex differences (2019): 49
SA-β-Galactosidase-Based Screening Assay for the Identification of Senotherapeutic Drugs.
Authors: Fuhrmann-Stroissnigg, Heike and Santiago, Fernando E and Grassi, Diego and Ling, YuanYuan and Niedernhofer, Laura J and Robbins, Paul D
Journal: Journal of visualized experiments : JoVE (2019)
Cellular and cytoskeletal alterations of scleral fibroblasts in response to glucocorticoid steroids.
Authors: Bogarin, Thania and Saraswathy, Sindhu and Akiyama, Goichi and Xie, Xiaobin and Weinreb, Robert N and Zheng, Jie and Huang, Alex S
Journal: Experimental eye research (2019): 107774
Tumor cell escape from therapy-induced senescence.
Authors: Saleh, Tareq and Tyutyunyk-Massey, Liliya and Murray, Graeme F and Alotaibi, Moureq R and Kawale, Ajinkya S and Elsayed, Zeinab and Henderson, Scott C and Yakovlev, Vasily and Elmore, Lynne W and Toor, Amir and Harada, Hisashi and Reed, Jason and Landry, Joseph W and Gewirtz, David A
Journal: Biochemical pharmacology (2019): 202-212
Page updated on December 6, 2024

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Catalog Number14030
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Physical properties

Molecular weight

494.50

Solvent

DMSO

Storage, safety and handling

H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22

Storage

Freeze (< -15 °C); Minimize light exposure
UNSPSC12171501

Platform

Flow cytometer

Excitation488 nm laser
Emission530, 30 nm filter
Instrument specification(s)FITC channel

Fluorescence microscope

ExcitationFITC filter set
EmissionFITC filter set
Recommended plateBlack wall, clear bottom
Expression of &beta;-gal was measured with Xite&trade; Green beta-D-galactopyranoside. 9L-LacZ cells (cells that overexpressed &beta;-gal) were incubated with Xite&trade; Green beta-D-galactopyranoside or FDG for 30 mins at 37 &deg;C. The signal was acquired&nbsp;with FITC channel using a NovoCyte Flow Cytometer (ACEA Biosciences).
Expression of &beta;-gal was measured with Xite&trade; Green beta-D-galactopyranoside. 9L-LacZ cells (cells that overexpressed &beta;-gal) were incubated with Xite&trade; Green beta-D-galactopyranoside or FDG for 30 mins at 37 &deg;C. The signal was acquired&nbsp;with FITC channel using a NovoCyte Flow Cytometer (ACEA Biosciences).
Expression of &beta;-gal was measured with Xite&trade; Green beta-D-galactopyranoside. 9L-LacZ cells (cells that overexpressed &beta;-gal) were incubated with Xite&trade; Green beta-D-galactopyranoside or FDG for 30 mins at 37 &deg;C. The signal was acquired&nbsp;with FITC channel using a NovoCyte Flow Cytometer (ACEA Biosciences).