Evaluation of the Pig-a, micronucleus, and comet assay endpoints in a 28-day study with ethyl methanesulfonate.

Ethyl methanesulfonate (EMS) was evaluated as part of the validation effort for the rat Pig-a mutation assay and compared with other well-established in vivo genotoxicity endpoints. Male Sprague-Dawley (SD) rats were given a daily dose of 0, 6.25, 12.

Report on stage III Pig-a mutation assays using N-ethyl-N-nitrosourea-comparison with other in vivo genotoxicity endpoints.

N-Ethyl-N-nitrosourea (ENU) was evaluated as part of the Stage III trial for the rat Pig-a gene mutation assay. Groups of six- to eight-week-old male Sprague Dawley (SD) or Fischer 344 (F344) rats were given 28 daily doses of the phosphate buffered saline vehicle, or 2.5, 5, or 10 mg/kg ENU, and evaluated for a variety of genotoxicity endpoints in peripheral blood, spleen, liver, and colon.

International Pig-a gene mutation assay trial: evaluation of transferability across 14 laboratories.

A collaborative international trial was conducted to evaluate the reproducibility and transferability of an in vivo mutation assay based on the enumeration of CD59-negative rat erythrocytes, a phenotype that is indicative of Pig-a gene mutation. Fourteen laboratories participated in this study, where anti-CD59-PE, SYTO 13 dye, and flow cytometry were used to determine the frequency of CD59-negative erythrocytes (RBC(CD59-)) and CD59-negative reticulocytes (RET(CD59-)). To provide samples with a range of mutant phenotype cell frequencies, male rats were exposed to N-ethyl-N-nitrosourea (ENU) via oral gavage for three consecutive days (Days 1-3).

Defining EMS and ENU dose-response relationships using the Pig-a mutation assay in rats.

In recent years, experimental evidence has accumulated that supports the existence of sublinear dose-response relationships at low doses of DNA reactive mutagens. However, creating the in vivo data necessary to allow for a more detailed dose-response modeling with the currently available tools might not always be practical. The purpose of the current work was to evaluate the utility of the Pig-a gene mutation assay to rapidly identify dose-response relationships for direct acting genotoxicants.

Evaluation of a modified CD71 MicroFlow method for the flow cytometric analysis of micronuclei in rat bone marrow erythrocytes.

The aim of this study was to evaluate a modified flow cytometric method for the quantification of micronuclei in rat bone marrow reticulocytes. The method identified uses the erythrocyte pure fraction from cellulose filtered bone marrow with slight modifications to the widely published MicroFlow(®) method developed by Litron Laboratories, Rochester, NY for the detection of micronuclei in peripheral blood. A number of experiments were conducted to compare the micronucleus induction measured by flow cytometry with traditional microscopic analysis in male rats treated daily for 2 days with appropriate vehicle controls or various doses of cyclophosphamide (CP), mitomycin C (MMC), vinblastine sulfate (VBS), 1,2-dimethylhydrazine (DMH), etoposide (ETO), colchicine (COL), or 4-nitroquinoline-1-oxide (4NQO).