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    ASTM E1439 - 12(2019)

    Standard Guide for Conducting the Frog Embryo Teratogenesis Assay-Xenopus (FETAX)

    Active Standard ASTM E1439 | Developed by Subcommittee: E50.47

    Book of Standards Volume: 11.06

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    Significance and Use

    5.1 FETAX is a rapid test for identifying potential developmental toxicity. Data may be extrapolated to other species including mammals. FETAX might be used to prioritize samples for further tests which use mammals. Validation studies using compounds with known mammalian or human developmental toxicity, or both, suggest that the predictive accuracy will exceed 85 % (2) . When evaluating a test material for mammalian developmental toxicity, FETAX must be used with and without a metabolic activation system (MAS). Use of this exogenous MAS should increase the predictive accuracy of the assay to approximately 95 %. The accuracy rate compares favorably with other currently available “ in vitro teratogenesis screening assays” (3). Any assay employing cells, parts of embryos, or whole embryos other than in vivo mammalian embryos is considered to be an in vitro assay.

    5.2 It is important to measure developmental toxicity because embryo mortality, malformation, and growth inhibition can often occur at concentrations far less than those required to affect adult organisms.

    5.3 Because of the sensitivity of embryonic and early life stages, FETAX provides information that might be useful in estimating the chronic toxicity of a test material to aquatic organisms.

    5.4 Results from FETAX might be useful when deriving water quality criteria for aquatic organisms (4).

    5.5 FETAX results might be useful for studying structure-activity relationships between test materials and for studying bioavailability.

    1. Scope

    1.1 This guide covers procedures for obtaining laboratory data concerning the developmental toxicity of a test material. The test utilizes embryos of the African clawed frog, Xenopus laevis and is called FETAX (Frog Embryo Teratogenesis Assay-Xenopus) (1).2 Some of these procedures will be useful for conducting developmental toxicity tests with other species of frogs although numerous modifications might be necessary. A list of alternative anurans is presented in Appendix X1.

    1.2 A renewal exposure regimen and the collection of the required mortality, malformation, and growth-inhibition data are described. Special needs or circumstances might require different types of exposure and data concerning other effects. Some of these modifications are listed in Appendix X2 although other modifications might also be necessary. Whenever these procedures are altered or other species used, the results of tests might not be comparable between modified and unmodified procedures. Any test that is conducted using modified procedures should be reported as having deviated from the guide.

    1.3 These procedures are applicable to all chemicals either individually or in formulations, commercial products or mixtures that can be measured accurately at the necessary concentrations in water. With appropriate modification these procedures can be used to conduct tests on the effects of temperature, dissolved oxygen, pH, physical agents, and on materials such as aqueous effluents (see Guide E1192), surface and ground waters, leachates, aqueous and solid phase extracts, and solid phase samples, such as soils and sediments, particulate matter, sediment, and whole bulk soils and sediment.

    1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.

    1.5 This guide is arranged as follows:



    Referenced Documents




    Summary of Guide


    Significance and Use


    Safety Precautions




    Water for Culturing Xenopus adults










    FETAX Solution Water






    Test Material




     Stock Solution


    Test Organisms














     Experimental Design


     Temperature and pH Requirements


     Beginning the Test




     Duration of Test


     Exogenous Metabolic Activation System (MAS)


     Biological Data


    Analytical Methodology


    Acceptability of the Test








     X1. List of Alternative Species

    Appendix X1

     X2. Additional Endpoints and Alternative Exposures

    Appendix X2

     X3. Concentration Steps for Range-Finding Tests

    Appendix X3

     X4. Microsome Isolation Reagents and NADPH Generating
       System Components,

    Appendix X4



    1.6 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

    2. Referenced Documents (purchase separately) The documents listed below are referenced within the subject standard but are not provided as part of the standard.

    ASTM Standards

    D1193 Specification for Reagent Water

    E729 Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and Amphibians

    E943 Terminology Relating to Biological Effects and Environmental Fate

    E1023 Guide for Assessing the Hazard of a Material to Aquatic Organisms and Their Uses

    E1192 Guide for Conducting Acute Toxicity Tests on Aqueous Ambient Samples and Effluents with Fishes, Macroinvertebrates, and Amphibians

    E1391 Guide for Collection, Storage, Characterization, and Manipulation of Sediments for Toxicological Testing and for Selection of Samplers Used to Collect Benthic Invertebrates

    E1525 Guide for Designing Biological Tests with Sediments

    E1706 Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates

    ICS Code

    ICS Number Code 07.080 (Biology. Botany. Zoology)

    UNSPSC Code

    UNSPSC Code 77101501(Risk or hazard assessment)

    Referencing This Standard
    Link Here
    Link to Active (This link will always route to the current Active version of the standard.)

    DOI: 10.1520/E1439-12R19

    Citation Format

    ASTM E1439-12(2019), Standard Guide for Conducting the Frog Embryo Teratogenesis Assay-Xenopus (FETAX), ASTM International, West Conshohocken, PA, 2019, www.astm.org

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