in vitro methods

/Tag:in vitro methods

Skin Sensitisation, Adverse Outcome Pathways and Alternatives

David Basketter

For toxicologists who are in any way associated with skin sensitisation, the last two decades have seen a series of fundamental changes. We have migrated from old-style guinea-pig assays, via the refined and reduced Local Lymph Node Assay (LLNA), to witness the imminent dominance of in vitro and in silico methods. Yet, over the same period, the use of the output data for human safety assurance has evolved from 'black box' risk assessment, via the quantitative risk assessment enabled by the LLNA measurement of potency, to a new period of relative uncertainty. This short review will endeavour to address these topics, all the while keeping a focus on three essential principles: a) that skin sensitisation potential is intrinsic in the molecular structure of the chemical; b) that test methods should have a mechanistic foundation; and finally c) that the only reason for undertaking any skin sensitisation work has to be the protection of human health.
You need to register (for free) to download this article. Please log in/register here.

The Use of Peptide Reactivity Assays for Skin Sensitisation Hazard Identification and Risk Assessment

G. Frank Gerberick

Over the past 20 years or more, investigators have been developing non-animal test methods for use in assessing the skin sensitisation potential of chemicals. In parallel with this effort, the key biological events of skin sensitisation have been well-characterised in an Adverse Outcome Pathway (AOP) proposed by the Organisation for Economic Co-operation and Development (OECD). The key molecular initiating event of this AOP is haptenation or covalent modification of epidermal proteins. In this review, the strengths and limitations of the Direct Peptide Reactivity Assay (DPRA) are described, and the more recently developed Peroxidase Peptide Reactivity Assay (PPRA). The DPRA has been formally validated and incorporated into an OECD Test Guideline (TG442C). The DPRA shows promise for assisting in hazard identification as well as for assessing skin sensitisation potency when used in an integrated testing strategy.
You need to register (for free) to download this article. Please log in/register here.

Genomics: An In Vitro Toxicology Point of View

Raffaella Corvi

Genomics, and in particular its derived discipline, toxicogenomics, are rapidly developing technologies, which permit studies on the impact of chemicals and drugs on gene expression in particular biological systems. Enormous amounts of data will be provided in the context of mechanistic and predictive toxicology from the use of the DNA microarray approach for the simultaneous analysis of the expression pattern of multiple genes. The high-throughput requirement of these approaches necessitates in vitrocell culture systems. This article will give a short overview of the areas of ECVAM's research in which this technology will initially be applied.
You need to register (for free) to download this article. Please log in/register here.

Combined In Vitro Tests as an Alternative to In Vivo Eye Irritation Tests

Yang Ying, Yang Xingfen, Zhang Wengai, Cai Jinheng, Xue Jinyu, Yang Guangyu, Tan Xiaohua, Xie Xiaoping, Xiong Xikun, Huang Junming and Guo Xiang

Accurate methods that test the eye irritation potential of chemicals, which do not involve the use of animals, are needed to meet new regulatory standards. We evaluated the applicability and predictive capacity of five in vitro tests for eye irritation: the Hen’s Egg Test-Chorioallantoic Membrane (HET-CAM) assay; the Chorioallantoic Membrane-Trypan Blue Staining (CAM-TBS) assay; the Fluorescein Leakage Test (FLT); the 3T3-Neutral Red Uptake (3T3-NRU) cytotoxicity assay; and the red blood cell (RBC) haemolysis assay. A panel of 16 chemicals (some at multiple concentrations) was assessed by using the five tests, and the results were compared with historical in vivo Draize test data. The results showed rank correlation and class concordance between the five alternative methods and the Draize test for the 16 chemicals. These in vitro assays had good predictive capacity, reproducibility and reliability when compared to the Draize test. The best relationship was between the HET-CAM, CAM-TBS and FLT results, and the modified maximum average score(s) (MMAS). A prediction model (PM) was developed, based on the maximum possible correlation between the MMAS and the HET-CAM, CAM-TBS and FLT results. The PM had a good predictive capacity when compared to the results of animal tests, indicating its potential value for the in vitro screening of chemicals for eye irritation effects.
You need to register (for free) to download this article. Please log in/register here.

Combined In Vitro Tests as an Alternative to In Vivo Eye Irritation Tests

Yang Ying, Yang Xingfen, Zhang Wengai, Cai Jinheng, Xue Jinyu, Yang Guangyu, Tan Xiaohua, Xie Xiaoping, Xiong Xikun, Huang Junming and Guo Xiang

Accurate methods that test the eye irritation potential of chemicals, which do not involve the use of animals, are needed to meet new regulatory standards. We evaluated the applicability and predictive capacity of five in vitro tests for eye irritation: the Hen’s Egg Test-Chorioallantoic Membrane (HET-CAM) assay; the Chorioallantoic Membrane-Trypan Blue Staining (CAM-TBS) assay; the Fluorescein Leakage Test (FLT); the 3T3-Neutral Red Uptake (3T3-NRU) cytotoxicity assay; and the red blood cell (RBC) haemolysis assay. A panel of 16 chemicals (some at multiple concentrations) was assessed by using the five tests, and the results were compared with historical in vivo Draize test data. The results showed rank correlation and class concordance between the five alternative methods and the Draize test for the 16 chemicals. These in vitro assays had good predictive capacity, reproducibility and reliability when compared to the Draize test. The best relationship was between the HET-CAM, CAM-TBS and FLT results, and the modified maximum average score(s) (MMAS). A prediction model (PM) was developed, based on the maximum possible correlation between the MMAS and the HET-CAM, CAM-TBS and FLT results. The PM had a good predictive capacity when compared to the results of animal tests, indicating its potential value for the in vitro screening of chemicals for eye irritation effects.
You need to register (for free) to download this article. Please log in/register here.

A Critical Evaluation of the 2011 ECHA Reports on Compliance with the REACH and CLP Regulations and on the Use of Alternatives to Testing on Animals for Compliance with the REACH Regulation

Horst Spielmann, Ursula G. Sauer and Ovanes Mekenyan

On 30 June 2011, the European Chemicals Agency published two reports, one on the functioning of the REACH system, the other on the use of alternatives to animal testing in compliance with that system. The data presented are based on information gained during the first registration period under the REACH system, which included high production volume chemicals and substances of very high concern, which have the most extensive information requirements. A total of 25,460 registration dossiers were received, covering 3,400 existing, so-called ‘phase-in’, substances, and 900 new, so-called ‘non-phase-in’, substances. Data sharing and the joint submission of data are reported to have worked successfully. In the registration dossiers for these substances, results from new animal tests were included for less than 1% of all the endpoints; testing proposals (required for ‘higher-tier’ information requirements) were submitted for 711 in vivo tests involving vertebrate animals. The registrants mainly used old, existing experimental data, or options for the adaptation (waiving) of information requirements, before collecting new information. For predicting substance toxicity, ‘read-across’ was the second most-used approach, followed by ‘weight-of-evidence’. In vitro toxicity tests played a minor role, and were only used when the respective test methods had gained the status of regulatory acceptance. All in all, a successful start to the REACH programme was reported, particularly since, in contrast to most predictions, it did not contribute to a significant increase in toxicity testing in animals.
You need to register (for free) to download this article. Please log in/register here.