Papers

Peer-reviewed International journal
Jul 17, 2019

The within‐ and between‐laboratory reproducibility and predictive capacity of the in chemico amino acid derivative reactivity assay: Results of validation study implemented in four participating laboratories

Journal of Applied Toxicology
  • Masaharu Fujita
  • Yusuke Yamamoto
  • Shinichi Watanabe
  • Tsunetsugu Sugawara
  • Koji Wakabayashi
  • Yu Tahara
  • Nobuyuki Horie
  • Keiichi Fujimoto
  • Kei Kusakari
  • Yoshihiko Kurokawa
  • Tsuyoshi Kawakami
  • Kohichi Kojima
  • Takashi Sozu
  • Takuto Nakayama
  • Takeru Kusao
  • Jon Richmond
  • Kleinstreuer Nicole
  • Bae‐Hwa Kim
  • Hajime Kojima
  • Toshihiko Kasahara
  • Atsushi Ono
  • Display all

Volume
42
Number
6
First page
1078
Last page
1090
Language
English
Publishing type
Research paper (scientific journal)
DOI
10.1002/jat.3834
Publisher
Wiley

The amino acid derivative reactivity assay (ADRA) is an in chemico alternative assay for skin sensitization listed in OECD test guideline 442C. ADRA evaluates the reactivity of sensitizers to proteins, which is key event 1 in the skin sensitization adverse outcome pathway. Although the current key event 1 evaluation method is a simple assay that evaluates nucleophile and test chemical reactivity, mixtures of unknown molecular weights cannot be evaluated because a constant molar ratio between the nucleophile and test chemical is necessary. In addition, because the nucleophile is quantified by HPLC, the frequency of co-eluting the test chemical and nucleophile increases when measuring multi-component mixtures. To solve these issues, test conditions have been developed using a 0.5 mg/mL test chemical solution and fluorescence-based detection. Since the practicality of these methods has not been substantiated, a validation test to confirm reproducibility was conducted in this study. The 10 proficiency substances listed in the ADRA guidelines were tested three times at five different laboratories. The results of both within- and between-laboratory reproducibility were 100%, and the results of ultraviolet- and fluorescence-based measurements were also consistent. In addition to the proficiency substances, a new positive control, squaric acid diethyl ester, was tested three times at the five laboratories. The results showed high reproducibility with N-(2-(1-naphthyl)acetyl)-l-cysteine depletion of 37%-52% and α-N-(2-(1-naphthyl)acetyl)-l-lysine depletion of 99%-100%. Thus, high reproducibility was confirmed in both evaluations of the 0.5 mg/mL test chemical and the fluorescence-based measurements, validating the practicability of these methods.

Link information
DOI
https://doi.org/10.1002/jat.3834
PubMed
https://www.ncbi.nlm.nih.gov/pubmed/35043433
URL
https://onlinelibrary.wiley.com/doi/pdf/10.1002/jat.3834
URL
https://onlinelibrary.wiley.com/doi/full-xml/10.1002/jat.3834
ID information
  • DOI : 10.1002/jat.3834
  • ISSN : 0260-437X
  • eISSN : 1099-1263
  • Pubmed ID : 35043433

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