Non Animal Testing, Alternative Test Methods, In Vitro Toxicology, IIVS | Sensitization
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Sensitization

Integrated Testing Strategies for Skin Sensitization

Determination of skin sensitization potential is a critical endpoint in the safety assessment of raw materials, chemicals, mixtures and formulations.  Although the Guinea Pig Maximization Test (GPMT) and Local Lymph node assay (LLNA) have historically been used to address this adverse effect, in vitro assays have been developed and validated in order to replace these resource-intensive animal tests.

Skin sensitization is the result of a series of biochemical “Key Events” (KEs) that involve covalent binding of (generally electrophilic) compounds to cellular proteins (KE1), activation of various pathways within skin cells (KE2) and priming of the immune system (KE3) that results in an allergic response upon repeat exposure to the substance.

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Direct Peptide Reactivity Activation Assay (DPRA, OECD 442C)

The DPRA  (OECD 442C) is an in chemico assay that  models the first key event in the skin sensitization Adverse Outcome Pathway (AOP) – skin, protein reactivity.

Compounds implicated in causing Allergic Contact Dermatitis (ACD) are generally electrophilic in nature. This assay identifies dermal sensitizers based on their reactivity with synthetic peptides containing the nucleophilic amino acid residues lysine and cysteine. Using LC/UV, conjugation of the test material with the peptides can be analyzed.

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ARE-Nrf2 Luciferase Keratinocyte Activation Test Method (OECD 442D)

KeratinoSensTM and LuSens are cell-based reporter gene assays that model the second key event in the Adverse Outcome Pathway for Dermal Sensitization, namely keratinocyte activation.  The assays measure the induction of a stably transfected luciferase gene under the control of the antioxidant response element (ARE) derived from the human AKR1C2 or rat NQO1 gene.

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Human Cell Line Activation Test (h-CLAT, OECD 442E)

The human Cell Line Activation Test (h-CLAT) is a cell-based assay that identifies skin sensitizers by examining changes in the expression of cell surface markers (CD54 and CD86) implicated in dendritic cell activation, the third key event of the skin sensitization AOP.  Following exposure of the THP-1 human monocyte cell line to the test substance, expression levels of CD54 and CD86 are quantified by flow cytometry and compared to controls.

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