Cell Senescence Assay

Cat. # CB011

(50 Tests in 35mm plate)

Description

Normal mammalian cells divide for a limited number of population doublings and eventually enter an arrested state in which the cells remain alive, but do not proliferate in response to mitogens, and assume a characteristic enlarged, flattened morphology. This process is senescence and thought to be a tumor suppressive mechanism and underlying cause of aging. Senescence-associated β-galactosidase (SA- β-gal) is a widely used biochemical maker for assessing senescence in cultured cells. The Cell Senescence Assay provide an easy-to-use method to detect SA-β-gal by staining cells with 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside (X-gal) at pH 6.0, a pH condition that suppress lysosomal β-galactosidase activity sufficiently to ensure that nonsenescent cells remain unstained.

Kit Components

100X Fixing Solution: One tube, 1.5 mL of 25% Glutaraldehyde

Staining Solution A: One tube, 1.5 mL of 500 mM Potassium Ferrocyanide

Staining Solution B: One tube, 1.5 mL of 500 mM Potassium Ferricyanide

Staining Solution C: Three tubes,1.5 ml of 1 M Citrate-Na₂HPO₄ Buffer, pH6.0, 50 mM MgCl₂

Staining Solution D: Two tubes – 2.0 mL of 5 M NaCl in each tube

X-gal Solution: Two tubes – 1.5 mL of 40 mg/mL X-gal in DMF in each tube

Materials Not Supplied

1. PBS

2. 37ºC Incubator

3. Light microscope

4. Senescent cells or tissue samples

*   Store X-gal solution protected from light at -20ºC. Store all other components at 4ºC. Crystal deposition, which comes from unreacted X-gal, may be observed after incubation of cells with working staining solution. It can be minimized by pre-filtering the working staining solution with a 0.2 µm filter.