Genetically akin wild-type and p21−/− changeable littermates, 12–18 weeks of age were acclimated for all abstracts (Kwon et al., 2002). Genomic DNA a and PCR were acclimated to affirm genotypes of the mice. Mice were housed in the aforementioned allowance beneath controlled conditions, including a 12 h light/dark cycle, with ad libitum admission to diet and water. Wild-type and p21-deficient animals accustomed injections of AOM in antiseptic acrid band-aid at the dosage of 10 mg/kg anatomy weight subcutaneously already a anniversary for 6 weeks. In parallel, ascendancy animals accustomed acrid injections. Animals were dead 4 weeks afterwards the aftermost AOM injection. For the apoptosis and admeasurement studies, animals were injected with 15 mg/kg AOM intraperitoneally (i.p.) and dead 6, 8, or 72 h later. Mice were injected i.p. with 50 μg of BrdU (Sigma, Saint Louis, MO, USA) per gram of mice anatomy weight 2 h afore killing.
Colons were removed, opened and bedfast amid two bottle slides and anchored in 70% ethanol. They were again decrepit with methylene dejected and the absolute cardinal of ACF were counted application transillumination and ablaze microscopy (Paulsen et al., 2001). ACF were beyond than adjoining accustomed crypts, with thickened epithelia and an adapted luminal aperture (Bird et al., 1989). Afterwards counting ACF, colons were disconnected into adjacent and distal portions and anchored
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