The assay is performed in the presence and the absence of a rat-liver homogenate activation system. In the absence of an activation system, 0.1 mI of a solution of the test sample and approximately 10e bacteria in 0.1 mI are added to 2 mi of top agar (0.6% ager, 0.6% NaCI, 0.05 mM L-histidine, 0.05 mM blotin). These components are mixed and poured on the surface of a plate containing 20 ml of Davis minimal agar. To treat in the presence of an activation system, 0.5 mi of 5-9 mix is added to the bacterla-test sample-top aGar mixture. S*9 is the 9,000 x g supernatant of liver hGECgOnBfe ifGo ratS Given 500 mg of ArocIor 12SO/kg five days before sacrifice. The S*9 mix contains per ml: 0.3 mI of S-9, 8 mM MgCI2, 33 mM KGI, 5 mM glucose-6-phosphate, 4 mM NADP and 100 nd4 sodium phosphate (pH 7.4). The S-9 mix is added to the bacteria, test sample and top agar. These components are mixed and immediately poured over the minimal ager plate. The reverted colonies are counted after the plates are Incubated at 37 deg. for 48 hr. Positive (known mutagens) and negative (solvent) controls are included in each assay. In assays in which an activaticr. system is present, an additional negative control (i.e., no S-9 mix) is included.