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KMID : 1177720170110010019
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Journal of Alternatives to Animal Experiments
2017 Volume.11 No. 1 p.19 ~ p.26
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Evaluation of the transferability of in vivo Pig-a gene mutation assay across three laboratories
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Chung Young-Shin
Park Bum-Soo
Han Se-Hee
Lee Ji-Yeon
Kim Ji-Young
Back Seng-Min
Park cho-Rong
Kim Su-Hwan
Lee Jong-Kwon
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Abstract
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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 RBCCD59- and RETCD59-, 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.
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KEYWORD
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Pig-a gene, CD59, in vivo mutagenecity, N-ethyl-N-nitrosourea, transferability
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