AN INTERNATIONAL EVALUATION OF THE MURINE LOCAL LYMPH-NODE ASSAY AND COMPARISON OF MODIFIED PROCEDURES

The murine local lymph node assay is a predictive test for the identif ication of skin-sensitizing chemicals. The method has been the subject both of national inter-laboratory studies and of extensive comparison s with guinea pig tests. In the investigations reported here, the loca l lymph node assay has been evaluated further in the context of an int ernational study comprising five independent laboratories. In addition , the influence of minor modifications to the standard assay procedure on the performance of the test has been examined. The modified proced ures investigated were exposure of mice for 4 rather than 3 consecutiv e days, excision of lymph nodes 4 rather than 5 days after the initiat ion of exposure and the use of an alternative isotope. All five labora tories, irrespective of whether the standard or a modified protocol wa s used, were able to identify accurately, and with comparable sensitiv ity, potassium dichromate and 2,4-dinitrochlorobenzene as skin sensiti zers. Using standard criteria, none of the laboratories recorded posit ive responses with methyl salicylate, a non-sensitizer. In the standar d protocol, lymph nodes are pooled for each experimental group and the vigor of responses measured as a stimulation index relative to vehicl e controls. A stimulation index of 3 or greater is considered to indic ate skin-sensitizing potential. One further modification adopted by th ree of the laboratories was to analyze nodes from individual animals a nd, thereby, permit statistical evaluation. This allowed a direct comp arison of statistical significance with the conventional stimulation i ndex as criteria for a positive response. The data indicate that, whil e statistical evaluation may provide, in some instances, for small inc reases in sensitivity, this may be at the expense of some loss of sele ctivity. There are, however, insufficient data presently to draw firm conclusions regarding the relative value of statistical analysis. Thes e studies demonstrate that the local lymph node assay is sufficiently robust to accommodate minor procedural and technical modifications wit hout material changes in test performance.