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We developed a new local lymph node assay (LLNA) that includes the elicitation phase termed LLNA:DAE for discrimination of borderline-positive chemicals as classified by the LLNA modified by Daicel based on ATP content (LLNA:DA) and for cross-sensitization testing. Although the LLNA:DA method could help identify skin sensitizers, some skin irritants classified as non-sensitizers by the LLNA were classified as borderline positive. In addition, the evaluation for the cross-sensitization potential between chemicals was impossible. In the LLNA:DAE procedure, test group of mice received four applications of chemicals on the dorsum of the right ear for induction and one application on the dorsum of the left ear for elicitation. Control group of mice received one chemical application on the dorsum of the left ear. We evaluated the sensitizing potential by comparing the weights of the lymph nodes from the left ears between the two groups. The results of using the LLNA:DAE method to examine 24 chemicals, which contained borderline-positive chemicals, were consistent with those from the LLNA method, except for nickel chloride (NiCl2). Two chemical pairs, 2,4-dinitrochlorobenzene (DNCB) with 2,4-dinitrofluorobenzene (DNFB) and hydroquinone (HQ) with p-benzoquinone (p-BQ), showed clear cross-sensitization with each other, while another chemical pair, DNFB with hexylcinnamic aldehyde (HCA) did not. Taken together, our results suggest that the LLNA:DAE method is useful for discriminating borderline-positive chemicals and for determining chemical cross-sensitization.

journal of toxicological sciences

Development of LLNA:DAE: a new local lymph node assay that includes the elicitation phase, discriminates borderline-positive chemicals, and is useful for cross-sensitization testing