DPRA.

OECD TG 442C: Direct Peptide Reactivity Assay

Direct Peptide Reactivity Assay (DPRA) is an in chemico method that assesses the skin sensitization hazard of chemicals.

The assay evaluates a substance’s ability to react with synthetic model peptides, providing a binary classification that identifies test substances as either skin sensitizers or non‑sensitizers.

Targeting Key Event 1 of the skin sensitization AOP

DPRA is included in OECD Test Guideline 442C and addresses Key Event 1 (KE1) of the Adverse Outcome Pathway (AOP) for skin sensitization: Covalent binding to skin proteins.

Best suited for neat, straightforward organic chemicals

Available data* show that DPRA performs best for neat, soluble organic chemicals with direct protein reactivity, and may also be applicable to mixtures of known composition. Substances requiring metabolic activation (pro‑haptens) or containing metals fall outside the assay’s applicability domain.

*Reference: OECD TG 442C, Appendix I

A first-line KE1 assay supporting DASS under OECD TG 497

In regulatory context requiring multiple assays addressing different key events, DPRA serves as a first‑line KE1 method under under OECD TG 497.

It efficientily complements GARDskin (OECD TG 442E) and supports Defined Approaches for Skin Sensitization (DASS), including:

  • 2o3 Defined Approach: Combination of DPRA and GARD®skin for hazard identification.
  • Integrated testing strategy (ITS): Combination of DPRA, GARD®skin, and in silico methods to support hazard identification and GHS/CLP potency sub-categorization (1A/1B).
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At a glance

Test systemIn chemico.
ComplianceOECD TG 442C.
GLP-compliant laboratory.
What it measures Percent peptide depletion (cysteine and lysine).
ReadoutBinary prediction: Skin sensitizer or non-sensitizer.
Turnaround time2-4 weeks for standard studies.
Fast track available.

How it works

The Direct Peptide Reactivity Assay (DPRA) is an in chemico test method used to assess a chemical’s potential to cause skin sensitization. It does so by measuring the chemical’s ability to form covalent bonds with proteins, a key molecular initiating event in the skin sensitization process. This approach is based on the well‑established biological principle: most skin sensitisers are electrophilic substances that react with nucleophilic amino acid residues in skin proteins.

To model this process, test substances are incubated with two synthetic model peptides, each containing a single nucleophilic amino acid: cysteine or lysine. These amino acids represent the most common protein binding sites involved in skin sensitization. The peptides are exposed to the test substance for approximately 24 hours under controlled conditions, allowing covalent binding reactions to take place.

After incubation, the amount of remaining (non‑reacted) peptide is measured using high‑performance liquid chromatography (HPLC) with gradient elution and UV detection at 220 nm. The loss of peptide, expressed as percent peptide depletion, directly reflects the protein‑reactivity of the test substance.

Next, the mean depletion values from the cysteine and lysine peptides are calculated and evaluated using a defined prediction model. Based on established threshold values, each substance is assigned to one of four reactivity classes: no/minimal, low, moderate, or high reactivity. These classifications support the distinction between skin sensitizers and non‑sensitizers.

The results from DPRA are often interpreted as part of Defined Approaches or Integrated Testing Strategies under OECD TG 497, rather than used as a stand‑alone assessment.

For a complete description of the protocol and data interpretation, see OECD Test Guideline 442C.

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