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Table of Validated and Accepted Alternative Methods


Last Updated: November 15, 2009
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Validation & Regulatory Acceptance Status of Alternative Test Methods & Testing Strategies

Endpoint Method Name Test Type1 Endorsement of Scientific Validity Regulatory Acceptance
      Lead Authority Subsequent Endorsement(s) International (click here for approved OECD TGs and here for Draft TGs) National/ Regional (for methods not yet accepted internationally)
Acute aquatic toxicity Upper threshold concentration step-down approach

In vivo

ESAC (2006)

      
Acute mammalian toxicity (oral) Deletion of Acute Oral Toxicity Test, Lethal Dose (LD50)

In vivo

EU (2001)

  

OECD TG 401 deleted (2002)

  
  Acute toxic class method

In vivo

  

ESAC (2007)

OECD TG 423 (2001)

  
  Fixed dose procedure

In vivo

  

ESAC (2007)

OECD TG 420 (2001)

  
  Up-and-down procedure

In vivo

ICCVAM (2001)

ESAC (2007)

OECD TG 425 (2006)

  
  Normal human keratinocyte neutral red uptake (NHK NRU) assay

In vitro2

ICCVAM (2006)

  

Draft OECD TG

US agencies (2008)

  Balb/c 3T3 neutral red uptake assay

In vitro2

ICCVAM (2006)

  

Draft OECD TG

US agencies (2008)

Acute mammalian toxicity (inhalation) Acute toxic class method

In vivo

    

OECD TG 436

  
  Fixed concentration procedure

In vivo

    

Draft TG OECD 433

  
Chronic toxicity Ending 1-year dog studies of pesticides

In vivo

ESAC (2006)

    

Revised US EPA Pesticide Data Requirements

Dermal penetration In vitro skin absorption methods

In vitro

OECD Expert Group (2002)

  

OECD TG 428 (2004)

  
Endocrine mechanistic screens Androgen receptor binding assay (rat prostate)

In vitro

      

OPPTS TG 890.1150 (EPA, 2009)

  Aromatase inhibition assay (human recombinant)

In vitro

      

OPPTS TG 890.1200 (EPA, 2009)

  ER-alpha transcriptional activation assay for estrogen agnoists3

In vitro

    

OECD TG 455 (2009)

  
  Estrogen receptor binding assay

In vitro

      

OPPTS TG 890.1250 (EPA, 2009)

  Steroidogenesis (H295R human cell line)

In vitro

      

OPPTS TG 890.1550 (EPA, 2009)

  US EPA Tier 1 Screening Battery

In vitro/In vivo

      

US EPA (2009)

Genotoxicity Bacterial reverse mutation (Ames) test

In vitro

    

OECD TG 471 (1997)

  
  In vitro cell gene mutation test

In vitro

    

OECD TG 476 (1997)

  
  In vitro chromosomal aberration test

In vitro

    

OECD TG 473 (1997)

  
  In vitro micronucleus test

In vitro

ESAC (2006)

  

Draft OECD TG 487

REACH Regulation

  In vitro sister chromatid exchange test

In vitro

    

OECD TG 479 (1986)

  
  In vitro unscheduled DNA synthesis test

In vitro

    

OECD TG 482 (1986)

  
  Saccharomyces cerevisiae gene mutation assay

In vitro

    

OECD TG 480 (1986)

  
  Saacharomyces cerevisiae mitotic recombination assay

In vitro

    

OECD TG 481 (1986)

  
Hematotoxicity: acute neutropenia Colony forming unit granulocyte macrophage (CFU-GM) assay

In vitro

ESAC (2006)

      
Ocular irritation Cytotoxicity/cell-function based in vitro assays: Cytosensor Microphysiometer modified and Fluorescein Leakage

In vitro

ESAC (2009)

      
Ocular corrosion Bovine corneal opacity permeability (BCOP) test

In vitro

ICCVAM (2007)

ESAC (2007)

OECD TG 437 (2009)

OECD Proficiency Standards

  
  Isolated chicken eye (ICE) test

In vitro or Ex vivo

ICCVAM (2007)

ESAC (2007)

OECD TG 438 (2009)

OECD Proficiency Standards

  
  Hen's egg test-chorioallantoic membrane (HET-CAM)

In vitro

      

EU Competent Authorities for Dangerous Substances Directive

  Isolated rabbit eye test

In vitro or Ex vivo

      

EU Competent Authorities for Dangerous Substances Directive

Phototoxicity 3T3 Neutral Red Uptake Phototoxicity Test

In vitro

ESAC (1997)

  

OECD TG 432 (2004)

  
  3T3 NRU Phototoxicity Test: Application to UV filter chemicals

In vitro

ESAC (1998)

  

OECD TG 432 (2004)

  
Pyrogenicity Human whole blood IL-1

In vitro

ESAC (2006)

ICCVAM (2008)4

  

European Pharmacopeia; US agencies

  Human whole blood IL-6

In vitro

ESAC (2006)

ICCVAM (2008)4

  

European Pharmacopeia; US agencies

  Human cryopreserved whole blood IL-1

In vitro

ESAC (2006)

ICCVAM (2008)4

  

European Pharmacopeia; US agencies

  PBMC IL-6

In vitro

ESAC (2006)

ICCVAM (2008)4

  

European Pharmacopeia; US agencies

  MM6 IL-6

In vitro

ESAC (2006)

ICCVAM (2008)4

  

European Pharmacopeia; US agencies

  Limulus amebocyte lysate (LAL) test

In vitro

      

EDQM/European Pharmacopeia (5.0); US Pharmacopeia (85)

Reproductive & developmental toxicity Embryonic stem cell test

In vitro

ESAC (2002)

      
  Micromass assay

Ex vivo

ESAC (2002)

      
  Whole rat embryo assay

Ex vivo

ESAC (2002)

      
Skin corrosion EST-1000 human reconstructed epidermis

In vitro

ESAC (2009)

  

OECD TG 431 (2004)

  
  Corrositex®

In vitro

ICCVAM (1999)

ESAC (2000)

OECD TG 435 (2006)

  
  EpiSkin®

In vitro

ESAC (1998)

ICCVAM (2002)

OECD TG 431 (2004)

Draft Revised TG 431

  
  EpiDermTM

In vitro

ESAC (1998)

ICCVAM (2002)

OECD TG 431 (2004)

Draft Revised TG 431

  
  Rat skin transcutaneous electrical resistance (TER)assay

Ex vivo

ESAC (1998)

ICCVAM (2002)

OECD TG 430 (2004)

Draft Revised TG 430

  
  SkinEthicTM human skin model

In vitro

ESAC (2006)

  

OECD TG 431 (2004)

Draft Revised TG 431

  
Skin irritation EpiSkin® skin irritation test

In vitro

ESAC (2007)

  

Draft OECD TG

EU test method B.46 in COM regulation 440/2008/EC

  EpiDerm® skin irritation test

In vitro

ESAC (2007)

  

Draft OECD TG

EU test method B.46 in COM regulation 440/2008/EC

  EpiDermTM SIT model

In vitro

ESAC (2008)

  

Draft OECD TG

EU test method B.46 in COM regulation 440/2008/EC

  SkinEthic RHE model

In vitro

ESAC (2008)

  

Draft OECD TG

EU test method B.46 in COM regulation 440/2008/EC

Skin sensitization Local lymph node assay (LLNA)

In vivo

ICCVAM (1999)

ESAC (1999)

OECD TG 429 (2002)

Draft Revised TG 429

  
  Reduced LLNA

In vivo

ESAC (2007)

ICCVAM (2009)

Performance Standards (2009)

    
  Nonradiolabelled LLNA: DA

In vivo

ICCVAM (2009)5

  

Draft OECD TG

  
  LLNA: BrdU-ELISA

In vivo

ICCVAM (2009)5

  

Draft OECD TG

  
Vaccine potency ELISA for erysipelas vaccines batch potency testing

In vitro

ESAC (2002)

    

EDQM/European Pharmacopeia

  ELISA for human tetanus vaccines batch potency testing

In vitro

ESAC (2000)

    

EDQM/European Pharmacopeia

  Toxin binding inhibition test for human tetanus vaccines batch potency testing

In vitro

ESAC (2000)

    

EDQM/European Pharmacopeia

1 All in vitro and ex vivo methods listed; in vivo methods proposed to reduce or refine animal use also listed
2 Replaces animal use for initial dose setting, but in vivo test required to complete assessment
3 TA assay is in process of being formally validated, but included here because of recent OECD TG
4 Subject to product-specific validation to demonstrate equivalence to the rabbit pyrogen test (RPT)
5 ICCVAM recommendations in stages of being finalized


Supplemental Information

A. Summary of test methods evaluated by ESAC and accepted by EU regulatory authorities

B. Summary of test methods evaluated by ICCVAM and accepted by US regulatory authorities

C. OECD Test Guidelines (TGs), Guidance Documents, Draft Test Guidelines

D. ICH test guidelines

E. VICH test guidelines