Test No. 442A: Skin Sensitization
Local Lymph Node Assay: DA
The Local Lymph Node Assay: DA (LLNA: DA) is a non-radioactive modification to the
LLNA method for identifying potential skin sensitizing test substances and measuring
the proliferation of lymphocytes they induce in the auricular lymph nodes. The method,
described in mouse (CBA/J strain), is based on measurement of the adenosine triphosphate
(ATP) content by bioluminescence as an indicator of this proliferation. A minimum
of four animals is used per dose group, with a minimum of three concentrations of
the test substance, plus a concurrent negative control group and, if appropriate,
a positive control group. The experimental schedule is during 8 days. The time from
animal sacrifice to measurement of ATP should not exceed 30 min. The procedure from
excision of lymph nodes to ATP measurement should be kept uniform for each animal
and completed within 20 minutes. The luciferin/luciferase method is applied to measure
the bioluminescence in Relative Luminescence Units (RLU). This study includes: measurements
(weighing, RLU), and clinical daily observations. The results are expressed as the
Stimulation Index (SI) obtained by calculation. The SI should be ¡Ý1.8 before further
evaluation of the test material as a potential skin sensitizer is warranted.
Published on July 23, 2010Also available in: French
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