생체 내 Pig-a 유전자 돌병변이시험에 대한 세 시험실의 전수가능성 평가
Evaluation of the transferability of in vivo Pig-a gene mutation assay across three laboratories
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Abstract
In order to establish a pig-a gene mutation assay as a regulatory test for evaluating in vivo genotoxicity in Korea, the transferability study as a stage II validation was performed using the test substance, N-ethyl-N-nitrosourea (ENU) across three laboratories. The lead laboratory (KIT) standardized the protocol using Rat MutaFlow® Kit and transferred the methodology to two participating laboratories (Hoseo University and Biotoxtech Co. Ltd.). Sprague Dawley rats were treated with vehicle or three dose levels of ENU (10, 20, 40mg/kg) by oral gavages on the 3 consecutive days. On the day 15 after the first treatment, bloods were collected and analyzed to estimate the percentage of reticulocytes (%RETs), mutant type of RETs and erythrocytes (RBC) using flowcytometer. In results, the exposure of ENU induced the dose-dependent increases in the mutant frequencies of RBC CD59- and RET CD59- , while %RETs were not affected. Data comparisons among three participating laboratories indicated the good transferability with the high correlation coefficients (0.97-0.99) between the lead and participating laboratories. In conclusion, the pig-a gene mutation assay may be the effective test for the estimation of in vivo gene mutation and the validation studies may be needed using more test substances in Korea.
참고문헌 (30)
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