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Stewart Laboratory Standard Operating Procedure

Stewart Laboratory Standard Operating Procedure               

Senescent-associated b-Gal Assay

  1. Wash cells with PBS
  2. Fix cells in 0.2% glutaraldehyde for 5’
  • dilute this in PBS
    •
    1. Wash cell with PBS
    2. Stain with X-gal solution (ingredients listed below) for 6 to 24 hours at 37°C

    Chemical Conc.

    		 Stocks Amount needed for 10 plates (40 ml)
    1 mg/ml X-gal 40 mg/ml  1 ml
    150 mM NaCl 5 M 1.2 ml
    2 mM MgCl2 1M 80 μl
    5 mM K3Fe(CN)6 100 mM  2 ml
    5 mM K4Fe(CN)6 100 mM 2 ml
    40 mM NaPi, pH 6.0  0.1M 16 ml

    17.72 ml dd H2O

    NaPi - 0.1M at pH 6.0