Animal and non-animal tests for cosmetics

The range of extremely cruel and painful animal tests are common to assess the alleged safety of cosmetic ingredients. Numerous in vitro tests have already been developed and accepted on a regulatory level.

Animal testing for cosmetics is prohibited in the EU, but there are still many countries (e.g. China) in which cosmetics testing on animals is still a common practice.

The following list gives a brief overview of the options available today. However, Doctors Against Animal Experiments are opposed to the use of animals, animal-derived materials (e.g. animal cells) and eggs.

Acute and chronic toxicity

3t3 nru test

Animal experiment: Rats or mice are force-fed a substance via gavage (feeding tube) once or multiple times over a longer period. Depending on the type and amount of the substance administered, the animals may suffer from diarrhea, fever or paralysis.

In vitro: The toxicity of cosmetic and other chemical products can be investigated by using human or other mammalian cell cultures. The tests are very sensitive and the cells die when exposed to toxic substances.

The 3T3 neutral red test is based on the principle that cells of a certain permanent mouse cell line are no longer able to take up a red dye after being exposed to a harmful compound.

Examples: 3T3 NRU test (Neutral Red Uptake Cytotoxicity Assay) 

Skin irritation test

kaninchen onevoice

Animal experiment: The test substance is applied onto the shaved skin of rabbits. An irritating substance can cause painful skin inflammations.

In vitro: The substance is analysed by applaing it to isolated human skin or skin cell cultures. Various test systems of human 3D skin models are commercially available which comprise the physiological properties of human skin.

Examples: EpiSkin, EpiDerm

Eye irritation test (Draize test)


Animal experiment: The test substance is applied into the eyes of rabbits. Depending on the type and dosage of the substance, painful inflammation and severe eye corrosion can occur.

In vitro: The HET-CAM test uses the membrane that lies directly under the shell of an incubated chicken egg, which contains veins and arteries but no nerves. The test substance is dripped onto this membrane and the resulting reaction is examined. Another method uses human skin cells.

Examples: EpiOcular ™ Human Cell Construct EIT

Skin allergy test

Animal experiment: The test substance is injected into the skin of guinea pigs to stimulate the animals’ immune system. If the test substance triggers allergic reactions after repeated contact, painful skin infections occur.

In vitro: It is possible to identify potential allergens using a chemical reaction (DPRA). Furthermore, there are (human) cell culture systems, used to test whether the addition of a test substance triggers intracellular signaling pathways related to allergy reactions.

Examples: DPRA (Direct Peptide Reactivity Assay), KeratinoSens Assay 

Phototoxicity (Harmful effects of sunlight)

Animal test: The test is carried out on rats or guinea pigs and is very similar to the skin allergy test. The test substance is injected into the animals’ skin. During the subsequent irradiation with UV A light, the animals are immobilised in tight plastic tubes for hours.

In vitro: The 3T3 neutral red test is based on the principle that cells of a certain permanent mouse cell line are no longer able to take up a red dye after the application of a harmful substance and UV light radiation. 

Skin absorption test (toxicokinetics)

Animal experiment: The test substance is applied on the skin of rats. The animals are kept in isolation in so-called metabolic cages. Their urine, feces and sometimes also blood samples are examined for the presence of the test substance.

In vitro: The test can be carried out on human skin samples. 

Damage to foetus or embryo (teratogenicity)

Animal experiment: The test substance is administered to pregnant rats or rabbits. The animals are killed at various time points during pregnancy to assess whether the substance is damaging to the mother and/or affects the development of the embryos. Teratogenic substances lead to malformations or stillbirths.

In vitro: There are numerous in vitro tests, for example with isolated embryos from mice, rats or rabbits (embryo culture), with permanent mouse cell lines (embryonic stem cell test) or with cells from mouse embryos (Limb Bud Micromass test).


Animal experiment: The test causes enormous suffering as the test substance is given to rats or mice every day throughout their life, e.g. force-fed through a gavage. It is analysed whether the animals develop tumors.

In vitro: Cell cultures from hamster embryos or mouse cell lines are used for the so-called transformation test. Human liver and skin cells are also suitable. 

Examples: Bhas CTA, Three CTA

Author: Julia Radzwill, biologist
Translation: Dilyana Filipova, PhD
11 August 2020


EU Science Hub. Validated test methods [retrieved 24.03.2020]
Cyprotex. In vitroHCS Manual (PDF) [retrieved 24.03.2020] 

Further Information

Animal testing-free cosmetics >>

The long road to the sales ban on animal testing for cosmetics >>