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Understanding In Vitro and In Vivo Toxicology Testing for Chemicals

Little Pro on 2018-02-11 Views:  Update:2018-10-17

Both in vitro and in vivo methods are routinely used by industries and regulatory authority in toxicity testing, GHS classification and chemical risk assessment. In this article, we will summarize the definition of in vitro and in vivo tests and provide a list of common validated in vitro toxicology test methods for chemicals. In the end we will explain some factors to consider when using in vitro toxicology methods for regulatory work.

Definition and Overview of In Vitro and In Vivo Test

An in vitro (latin: "in the glass") test is a study that is performed outside of a living organism (i.e, glass, petri dishes). The test usually involves the use of isolated tissues, organs or cells. In contrast, an in vivo (latin: "in the living") test is a study that is performed in living organisms (i.e, rat, rabbit).

Both in vitro and in vivo methods can be used to predict the inherent hazard properties of chemical substances. However, results obtained from in vitro studies cannot often be used directly to predict biological responses of organisms to chemical exposure in vivo.

That does not mean that in vitro data is of less value. In fact, in vitro test is gaining more attention in regulatory community since it can reduce the number of animals used significantly while offering reliable predictions for some toxicological endpoints. Take EU REACH for example, an in vivo eye irritation or skin irritation study shall only be considered in case the in vitro tests are not applicable for a substance or the results obtained are not adequate for classification and risk assessment.

List of Common Validated In Vitro Toxicology Methods

The table below summarizes a list of common validated in vitro toxicology testing methods for chemicals. In vitro methods are mainly used for skin/eye irritation test, skin sensitisation test and genotoxicity tests. They cannot replace acute toxicity, repeated dose toxicity, development and reproductive toxicity, and carcinogenicity yet.

Endpoints In vitro methods
Skin Irritation
  • OECD TG 430 In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test
  • OECD TG 431 In Vitro Skin Corrosion: Human Skin Model Test
  • OECD TG 432 In Vitro 3T3 NRU Phototoxicity
  • OECD TG 435 In Vitro Membrane Barrier Test Method for Skin Corrosion
  • OECD 439 In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method

Above methods may be used to replace the following traditional in vivo test.

  • OECD TG 404 Acute Dermal Irritation / Corrosion 
Serious Eye Damage/Eye Irritation
  • OECD TG 437 In Vitro Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants
  • OECD TG 438 In Vitro Isolated Chicken Eye Test Method for Identifying Ocular Corrosives and Severe Irritants
  • OECD TG 460 Fluorescein Leakage Test Method (FL)
  • OECD TG 491 Short Time Exposure Test Method (STE)
  • OECD TG 492 Reconstructed human Cornea-like Epithelium Test Method (RhCE) 

Above methods may be used to replace the following traditional in vivo test.

  • OECD TG 405 Acute Eye Irritation / Corrosion 
Sensitisation
  • OECD TG 442D In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method
  • OECD TG xxxx In Vitro Skin Sensitisation: Human Cell Line Activation Test (h-CLAT)

Any method above alone cannot replace the following traditional in vivo test.

  • OECD TG 429 Skin sensitisation: Local Lymph Node Assay (LLNA) - preferred by REACH
  • OECD TG 406 Skin sensitisation: the guinea pig maximisation test (GPMT) and the Buehler test 
Genotoxicity

The following in vitro studies are usually conducted first. If there is any positive response in the in vitro study, a follow-up in vivo study for the same endpoint is usually required.

  • OECD TG 471 Bacterial reverse mutation test (Ames test)
  • OECD TG 473 In vitro mammalian chromosome aberration test 
  • OECD TG 476 In vitro mammalian cell gene mutation test
  • OECD TG 482 DNA Damage and Repair, Unscheduled DNA Synthesis in Mammalian Cells In Vitro

Factors to Consider When Using In Vitro Methods

There are three factors to consider when using in vitro methods for regulatory work. Firstly, we need to know whether in vitro methods are accepted by regulatory authority in your jurisdiction or not. Some countries do not accept in vitro data  at all while some countries/industries ban in vivo animal testing completely. Secondly, we need to consider animal welfare issue. In vitro methods should be used more to provide screening info for in vivo testing. Thirdly, we need to know the limitations of in vitro methods. For example, OECD TG 437 The Bovine Corneal Opacity and Permeability Test Method (BCOP) is an in vitro assay that can be used to identify chemicals (substances or mixtures) as either i) causing “serious eye damage” (Cat 1 ), or ii) not requiring classification at all. However, it cannot be used to determine if a chemical substance is an eye irritant or not (Cat 2). 

References

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 Tags: Topics - CRAToxicology and Health Risk Assessment

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