FAQ: 96-Well Cellular Senescence Activity Assay

Q: How is SA-ß-Gal activity measured with this assay?

A: This assay uses a fluorometric substrate to detect SA-ß-Gal enzyme activity in cell lysates.  The buffers included in this kit maintain pH 6.0 throughout the assay to ensure optimal conditions for measuring SA-ß-Gal while suppressing lysosomal ß-gal activity. 


Q: What is the detection mechanism?

A: Senescence is  measured by detecting SA-ß-galactosidase activity in cell lysates.  A non-fluorescent substrate is hydrolyzed by SA-ß-galactosidase to emit a blue fluorescence, which is read at 360/465nm.  The stop solution that is added to the reaction contains components that will maximize the fluorescence while stopping the reaction at a fixed time point.


Q: Can collagen coated plates be used with this assay?

A: Collagen will not interfere with this assay and it is fine to grow cells on a collagen coated plate.


Q: Is this assay compatible with any cell line?

A: The senescence-associated ß-galactosidase (SA-ß-Gal) activity is a universal marker for any senescence cell and this assay is not cell type specific.


Q: Can tissues be used with this assay?

A: Tissues can be used with this assay; they will need to be resuspended in the 1X lysis buffer included with the kit and either sonicated or homogenized before using in the assay.  RIPA buffer is not compatible with the substrate, which is pH dependent, and should not be used in place of the 1X lysis buffer.


Q: How many cells should be used with this assay?

A: The cell number used with this assay can vary depending on the percentage of cells expected to be senescent, but a good starting point is 20,000 to 50,000 cells per well. If you find that you have a large percentage of senescent cells in your population, you may need to dilute from there.