Eye And Skin Safety
For over 50 years, MB Research Laboratories has provided eye and dermal hazard identification services to the specialty chemical, agrichemical, pharmaceutical, and consumer product industries.
Since 1989, MB Research has been leading the development and promotion of alternative and in vitro assays, inventing, adopting, and adapting non-animal replacements. Modernize Today!
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Eye Safety
In Vitro Assays
The Irritection® Assay System uses an in vitro method to determine ocular irritation and predict U.N. GHS classification for chemicals or mixtures. This study is designed to comply with the standards set forth in the OECD Guideline for the Testing of Chemicals No. 496.
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The purpose of this study is to provide classification of chemicals concerning their eye irritation potential using an alternative to the Draize Rabbit Eye Test, according to the OECD Test Guideline No. 492, “Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage”. The EpiOcular™ EIT is intended to differentiate those materials that are UN GHS No Category (i.e., do not meet the requirements for UN GHS classification) from those that would require labeling as either UN GHS Category 1 or 2.
Alternative Assays
The Bovine Corneal Opacity and Permeability (BCOP) assay is used to evaluate the eye irritation potential of chemicals, formulations, and products. Developed by Gautheron et al. (1992), this assay uses fresh bovine corneal tissue sourced as a by-product from abattoirs, offering an alternative to traditional in vivo testing.
The BCOP assay measures two key indicators of ocular damage:
Opacity, indicating structural damage or stromal swelling due to protein denaturation.
Permeability, reflecting the disruption of corneal barrier function, often caused by compromised cell membranes.
Both measurements are used to calculate an In Vitro Irritation Score (IVIS), which is used to assign a classification category.
For enhanced evaluation, histopathological analysis can be included to assess the extent and depth of corneal injury. This includes analysis of swelling, hydration levels, and tissue morphology which can provide valuable insight into the nature and severity of the irritation.
The BCOP test is widely accepted by regulatory authorities and offers a rapid, reproducible, and ethical method for screening eye irritation potential in a wide range of products. The BCOP assay is internationally recognized and accepted under the following regulatory frameworks:
OECD Test Guideline 437 – for identifying substances that cause serious eye damage (UN GHS Category 1) or that do not require classification (No Category).
U.S. EPA – accepted as part of a weight-of-evidence approach for evaluating ocular irritation potential under the Toxic Substances Control Act (TSCA) and Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA).
EU REACH – utilized for classification and labeling under the European CLP Regulation in line with the Globally Harmonized System (GHS).
Health Canada and other international agencies – recognized for hazard identification and regulatory submissions involving cosmetic, chemical, agrochemical, and household product testing.
The BCOP test provides a rapid, reliable, and reproducible alternative to animal-based methods—delivering GLP-compliant data suitable for regulatory submission worldwide.
The objective of this study is to determine the potential for ocular irritation using OECD Guidelines for the Testing of Chemicals, Nos. 492 and 437, which serve as alternatives to the traditional Draize methodology. The use of two internationally-validated, OECD-accepted non-animal tests can provide a GHS categorization for the vast majority of test articles. The combination of the Bovine Corneal Opacity and Permeability Test and EpiOcular™ Eye Irritation Test followed by a Weight-of-Evidence analysis of the results allows for the classification of GHS Category 2.
Skin Safety
In Vitro Assays
Irritation
The Irritection® Assay System uses an in vitro method to determine dermal irritation and predict U.N. GHS classification for chemicals or mixtures.
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Corrosion
The Corrositex® assay is performed using a kit produced and distributed by In Vitro International. The kit contains tubes of proprietary buffers, a Chemical Detection System (CDS), and components used to make synthetic proteinaceous macromolecular bio‐barriers. After preliminary testing to determine if the test article is compatible with the Corrositex® assay (qualification) and to determine cut‐off times (categorization), the test article is applied topically on prepared bio‐barriers set atop vials of CDS. The amount of time it takes for the test article to penetrate the bio‐barriers and cause a visually detectable change in the CDS (breakthrough time) can be used to determine the UN Packing Group or the UN GHS subcategory (classification).
Sensitization
Upon exposure to skin sensitizers, the KeratinoSens™ Test measures activation of Keap1-Nrf2-antioxidant/electrophile response element (ARE). These tests use an immortalized, adherent, human keratinocyte cell line (HaCaT) that was transfected with a plasmid to monitor luciferase gene induction.
The DPRA is an in chemico method that quantifies cysteine‐ or lysine‐containing peptide depletion following 24 hours incubation with the test article (TA). Relative peptide concentration is measured by HPLC with gradient elution and UV detection at 220 nm. Cysteine and lysine peptide percent depletion values are then calculated and used in a prediction model, which allows assigning the TA to one of four reactivity classes used to support the discrimination between sensitizers and sonosensitizers.
Additional Assays
Irritation
This study is designed to evaluate local responses to extracts and complies with the standards set forth in the current International Organization for Standardization (ISO) International Standard 10993-23:2021 (E) Tests for Irritation.
Irritation/Corrosion
Sensitization
The Local Lymph Node Assay (LLNA) is alternative study to the Guinea Pig Sensitization Test used for determining the sensitizing potential of materials. Following exposure to a sensitizing test substance, lymphocyte proliferation occurs in the lymph node local to the site of exposure. The LLNA measures increased proliferation of lymphocytes in the auricular lymph node which drain the site of exposure; ears). Proliferation is assessed by determining the incorporation of the thymidine analog, bromodeoxyuridine (BrdU) into the DNA of lymph node cells using an enzyme-linked immunosorbent assay (ELISA).
Contact dermal sensitization is an immunological process where the host, through repeated skin exposure, acquires a specific allergic sensitivity to a substance. In the Buehler model, contact dermal sensitivity is manifested as increased erythema.
Non-Guideline Eye and Skin Safety Tests
In Vitro Eye Irritation
Alternative Eye Irritation
DB-ALM Protocol No. 96
The HET-CAM uses the highly vascularized chorioallantoic membrane (CAM) of a fertilized egg as a biological model for mucous membranes. Due to its sensitivity to chemical irritants, the CAM provides a reliable surrogate for evaluating eye irritation potential. Its inflammatory response closely resembles that of conjunctival tissue in the human eye.
HET-CAM is considered a valuable alternative to the traditional Draize Eye Test and aligns with the 3Rs principle (Replace, Reduce, and Refine). While HET-CAM alone may not fully satisfy regulatory testing requirements, it is a powerful tool for early-stage screening, formulation refinement, and reducing reliance on in vivo methods during product development.
The chorioallantoic membrane (CAM) of a fertilized chicken egg serves as an analog to the human conjunctiva with remarkable structural similarities in vasculature. Varying concentrations of a test article are administered to the CAMs of sets of eggs and qualitative measurements of vascular changes are performed in order to evaluate eye irritation potential.
This test is used to determine the potential for ocular irritation using an alternative to the Draize methodology. The methodology is based on that described in An Improved CAM Assay for Predicting Ocular Irritation Potential, Bagley, D.M., Rizvi, P.Y., Kong, B.M., and De Salva, S.J. (1988), Alternative Methods in Toxicology, Vol. 6, Progress in In vitro Toxicology, pp. 131-138, and "Bovine Corneal Opacity and Permeability Test: An In Vitro Assay of Ocular Irritancy,” (1994); Gautheron, Pierre; Dukic, Martine; Alix, Danielle and Sina, Joseph F.; Fundamental and Applied Toxicology 18, 442-449, and will include an analysis based on the current OECD Guideline for the Testing of Chemicals No. 437.
The PorCORA is an ex vivo alternative assay developed by MB Research that utilizes isolated, freshly harvested corneas from the eyes of swine in order to evaluate the corrosive potential of a test chemical by assessing:
(1) Sodium fluorescein stain retention, visualized using a transilluminator
(2) Time of sodium fluorescein clearance (up to 21 days post‐exposure)
The porcine corneas are maintained in cell culture medium with antibiotics for at least 21 days to model the Draize Rabbit Eye Test.
For the latest publication about PorCORA, read MB Research's article from Cutaneous and Ocular Toxicology 2024 Vol. 43.
In Vitro Skin Irritation
The EpiDerm™ MTT Viability Assay utilizes three-dimensional reconstructed human epidermis (RhE) tissues. Following exposure to a test article (TA) and incubation, tissue viability is measured via MTT reduction. The RhE tissues are composed of primary human cells, which have been cultured to form a stratified, highly differentiated squamous epithelium morphologically similar to that of human skin.
How can I reduce the cost of eye irritation, skin irritation, and skin sensitization testing?
Cost savings can be achieved by using validated in vitro (non-animal) test methods. These methods are generally faster and often eligible for regulatory acceptance by the EPA, FDA, and EU authorities. In addition, bundling multiple tests can reduce the total cost of the study by sharing controls.
How does selecting a GLP-compliant lab optimize long-term cost?
Working with a GLP-certified lab ensures data integrity and regulatory compliance, minimizing the risk of study repetition or delays in submission. GLP compliance is often a prerequisite for EPA, REACH, or FDA submissions, so investing in quality testing upfront reduces long-term regulatory setbacks.
