Local Lymph Node Assay (LLNA)

MDS offers a "Standard LLNA" (OECD TG429 and EPA OPPTS 870.2600). The Mouse Local Lymph Node Assay is accepted by the EPA, OECD, and FDA as the preferred alternative to the Guinea Pig Sensitization Test.

As a supplement testing, MDS also offers optional immunophenotypic marker evaluation by Flow Cytometry to further discriminate "true sensitizers" from "false-positive irritants” and enhance the results.

LLNA Options:

• Basic Dermal Irritation: Three concentrations 100%, 50%, 25% (for Non-Irritants) and two mice/concentration
• Quantitative Dermal Irritation: (Ear Swelling) Range-Finder six concentrations (e.g. 100%, 50%, 25%, 10%, 5%, 2.5%) two mice/concentration
• Modified OECD TG-429: Three concentrations of Test Article plus Vehicle Control group; Historical Positive Control) Pooled Nodes from each group. Four (4) mice/group
• Modified OECD TG429 does not include irritation/systemic toxicity Pre-Screen test.
• Standard EPA/OECD protocol+ Basic Irritation : (OECD TG-429 and OPPTS 870.2600 compliant. Includes Five (5) mice/group, Concurrent 25% Hexyl Cinnamic Aldehyde (HCA) Positive Control; Vehicle Control + 3 concentrations of Test Article, n=5/group) Basic Dermal Irritation/Toxicity Pre-Screen
• Regulatory EPA/OECD Protocol +Full Irritation/Toxicity. (OECD TG 429 and OPPTS 870.2600 compliant. Includes Quantitative Range Finding irritation screen + 3 concentrations per Test Article in main study with Positive and Vehicle Controls five (5) mice/group)
• Note: Although EPA/OECD Test Guidelines specify a minimum or 3 test concentrations, in the case of substances with dermal irritation properties, or in the absence of previous mouse ear dermal irritation data, the LLNA may require testing of 4-5 test concentrations to conduct a proper evaluation.
• Immunophenotyping: During the "induction phase" of sensitization, 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 or Iodine analog, bromodeoxyuridine (BrdU), into the DNA of lymph node cells using flow cytometric method. This "supplement testing " reduces background noise and false positives, by quantitatively assessing irritation, (via ear swelling) and by incorporating optional immunophenotype analysis to resolve and discriminate false-positive irritants %B220, CD3, CD4, CD8, I-Ak (MHC), CD69, CD25, CD44 + CD62lo+ lymph node cells