Biological activity: The material is tested for biological activity by conducting mutagenesis titration curves with at least one of the following three tests: benzo(a)pryrene, 2-anthramine, and 2-acetylaminofluorene.
Preparation: The material is prepared from 8-10 week old Sprague Dawley male rats. The animals are quarantined for two weeks prior to use. Preparation of S9 is by the procedure describes by Ames, et. Al. Induction: Animals induced with Aroclor 1254 are administered one intraperitoneal (i.p.) injection of Aroclor 1254 (500 mg/kg) suspended in corn oil five days prior to sacrifice, and starved 12 hours prior to sacrifice. Animals induced with 3-methylcholanthrene are administred three daily i.p. injections 24 hours after the last injection.
Rat Liver S9 Homogenate is used in Ames Mutagen Assay, to assess the mutagenic potential of chemical compounds. It is also used in assays that measure the metabolism of drugs and other xenobiotics. Many xenobiotics, neurotransmitters, steroids, and other hormones are metabolized by sulfate conjugation, a reaction catalyzed by the class of sulfotransferase enzymes. The common sulfate group donor is adenosine 3’-phosphate 5’-phosphosulfate.
Rat Liver S9 Homogenate is a Supernatant fraction obtained from an organ (usually liver) homogenate by centrifuging at 9000 g for 20 minutes in a suitable medium; this fraction contains cytosol and microsomes. The microsomes component of the S9 fraction contain cytochrome P450 isoforms (phase I metabolism) and other enzyme activities. The cytosolic portion contains the major part of the activities of transferases (phase II metabolism).
As required, sufficient S9 should be allowed to thaw at room temperature only until liquid, but still cold. Place the thawed S9 on ice. Discard any thawed, unused S9 at the end of the day. Although produced with aseptic technique, S9 cannot be sterilized by filtration or other means. Therefore, during use, it is imperative that S9 and S9 Mix be kept on ice at all times to prevent microbial growth and enzyme denaturation. If mixed with molten top agar for use in mutagenesis plates assays, the S9 Mix should be added immediately prior to pouring the agar plates. If kept at 45°C in the molten top agar for more than a few seconds, activity of the S9 Mix will be reduced.
Key Applications: Ames Mutagen Assay, Study of xenobiotic metabolism and drug interactions
Application Areas: Immunoassays
Product Type: Reagents
NOTES: Frozen liquid, ships on dry ice; Inducing Agent: Aroclor 1254
Storage & Handling: The S9 preparations are shipped in dry ice. Upon receipt, the S9 must be transferred to storage at -70°C or below.