Use of a B cell marker (B220) to discriminate between allergens and irritants in the local lymph node assay

G. Frank Gerberick, Lynn W. Cruse, Cindy A. Ryan, Ben C. Hulette, Joel G. Chaney, Robert A. Skinner, Rebecca J. Dearman, Ian Kimber

    Research output: Contribution to journalArticlepeer-review


    It has been shown that exposure of mice to contact allergens induces B cell activation in the draining lymph nodes (DLN), as seen by an increase in the percentage of B220+ or IgG/IgM+ cells. We have now examined whether the measurement of the percentage of B220+ cells could be used as an alternative or supplementary endpoint for the local lymph node assay (LLNA) to differentiate between allergenic responses and those few irritants that induce low-level proliferation in the DLN. Mice were treated on the ears, daily for 3 consecutive days, with various allergens (1-chloro-2,4-dinitrobenzene, α-hexylcinnamaldehyde, trinitrochlorobenzene, isoeugenol, and eugenol) or irritants (benzalkonium chloride, methyl salicylate, salicylic acid, and sodium lauryl sulfate). The DLN were excised 72 h following the final topical treatment, and the cells were prepared for B220 analysis using flow cytometry. The percentage of B220+ cells in lymph nodes derived from test and vehicle-treated animals was determined for 5 allergens and 4 irritants tested in multiple experiments (n = 3 to 17). As expected, the percentage of B220+ B cells was increased with each of the allergens tested, whereas irritant treatment did not cause similar increases. Moreover, the method was reproducible. For example, the strong allergen, 1-chloro-2,4-dinitrobenzene and the weak allergen, α-hexylcinnamaldehyde were identified as allergens in 17 of 17 and in 12 of 13 experiments, respectively. The percentage of B220 values for each chemical treatment (41 observations for allergens; 28 observations for irritants) versus the percentage of B220 values for the concurrent vehicle controls were plotted, and a classification tree model was developed that defined a B220 test: vehicle ratio cutoff of 1.25 for discriminating between allergens (>1.25) and irritants (
    Original languageEnglish
    Pages (from-to)420-428
    Number of pages8
    JournalToxicological Sciences
    Issue number2
    Publication statusPublished - Aug 2002


    • Allergens
    • B cells
    • Flow cytometry
    • Irritants
    • Local lymph node assay


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