Detection of tyrosinase-specific antibodies by ELISA. Representative results of three independent experiments for each patient--canine sera from patients A (panel a; 100 µg huTyr DNA dose), B (b; 100 µg huTyr DNA dose), and C (c; 1500 µg huTyr DNA dose)--were serially diluted at 1:20, 1:40, 1:80, 1:160, and 1:320, 1:640, and 1:1280. Each serum dilution was added in triplicate to individual wells (50 µl/well) containing recombinant huTyr (a-c) or myelin-basic protein, a nonspecific substrate (data not shown). The peak increase in tyrosinase-specific titer, compared to the preimmune serum, was detected for all patients at recheck 2: patient A at 1:1280, patient B at 1:640, and patient C at 1:320.