Skin / Dermal Sensitization

MB Research Labs has provided skin sensitization assessment services to the specialty chemical, agrichemical, pharmaceutical, consumer product, and medical device industries for 50 years. In addition to offering traditional testing methods, as new technology and knowledge of biology emerges, we invent, adopt, and adapt non-animal (in vitro and alternative) sensitization assays.

What is a Skin Sensitizer?

A skin sensitizer as defined by the Globally Harmonized System for Classification and Labelling of Chemicals (GHS) as “a substance that will lead to an allergic response following skin contact”. (UNECE, 2004)

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In Vitro Assays

KeratinoSens™
OECD 442D, ISO 10993-10

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.

Human Cell Line Activation test (h-CLAT)
OECD 442E/GHS, ISO 10993-10

To determine the skin sensitization potential of a test article due to cell activation by the measurement of changes in the expression of dendritic cell surface markers (CD86, CD54) via flow cytometry. The THP-1 human monocytic cell line serves as the test system. The h-CLAT is recommended by EURL ECVAM as part of an Integrated Approach to Testing and Assessment (IATA) to support the discrimination between sensitizers (i.e., UN GHS Category 1) and non-sensitizers for the purpose of hazard classification and labeling, per OECD Guideline 442E “In Vitro Skin Sensitization: Human Cell Line Activation Test (h-CLAT)”.

In Vitro Sensitization in EpiDerm™ (IVSA)
Non-Guideline

Developed and designed by MB Research Labs, the IVSA is an in vitro test that quantifies interleukin‐18 (IL‐18) release from keratinocytes following test article (TA) administration to reconstructed human epithelial (RhE) tissue. TIL‐18 is quantified via a commercial ELISA kit. IL‐18 release from TA‐treated tissues is compared that of vehicle‐treated controls. Solids, extracts, mixtures, insoluble chemicals and soluble chemicals are eligible for evaluation via the IVSA. Results are used to support the discrimination between sensitizers and non-sensitizers, and can be used for potency ranking.

Direct Peptide Reactivity Assay (DPRA)
OECD TG 442C

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.

Alternative Assays

Local Lymph Node Assay (LLNA) Flow Cytometry
OECD 429
To determine the sensitizing potential of topically applied test material utilizing the LLNA, and measurement of lymphocyte proliferation by BrdU incorporation (by flow cytometry). Our protocol is not a guideline assay but is designed to be compliant with OECD TG 429
Local Lymph Node Assay (LLNA-BrdU ELISA)
OECD 442, ISO 10993-10, OCSPP 870.2600
The local Lymph Node Assay (LLNA) is an alternative study to the Guinea Pig Sensitization Test, used for determining sensitization potential of materials. Following exposure to a sensitizing test substance, lymphocyte proliferation occurs in the local lymph node. The LLNA measures increased proliferation of lymphocytes in the auricular lymph nodes (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 flow cytometric method.

Additional Assays

Delayed Contact Dermal Sensitization Test – Buehler Method
OECD 406, ISO 10993-10, OSCPP 870.2600

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.

Guinea Pig Maximization Test (GMPT)
OECD 406, ISO 10993-10
Contact dermal sensitization is an immunological process where the host, through repeated skin exposure, acquires a specific allergic sensitivity to a substance. In the Guinea Pig Maximization (GPMT) model, contact dermal sensitivity is manifested as increased erythema.

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