Abstract: Objective To establish an immunological method for detecting antibodies of Penicillium marneffei. Methods The
recombinant Mp1p protein of Penicillium marneffei was expressed in Pichia pastoris and labeled with HRP (Mp1p-HRP) with a
modified sodium periodate method. A double-antigen sandwich enzyme-linked immunosorbant assay (ELISA) was
established by determining the optimal coating concentration of Mp1p protein and the concentration of the detecting protein
Mp1p-HRP. The sensitivity and specificity of the assay was evaluated by detecting Mp1p antibodies in 100 serum samples
from healthy donors, 15 samples from culture-confirmed penicilliosis patients, and 21 samples from patients with
culture-confirmed other fungal infections. Results A double-antigen sandwich ELISA was successfully established for
detecting Mp1p-specific antibody. The specificity of the assay was 100% (121/121) for detecting Mp1p-specific antibody in the
sera from healthy donors and patients with other fungal infection. The detection results of the 15 serum samples from patients
with culture-confirmed penicilliosis showed positivity for Mp1p antibody in 2 samples and Mp1p antigen positivity in 12
samples; combining the detection results of Mp1p antigen and antibody obviously increased the diagnostic sensitivity to 93.3%
(14/15). Conclusion The double-antigen sandwich ELISA shows a high specificity in detecting Mp1p-specific antibody, and
simultaneous detection of Mp1p antigen and antibody can increase the diagnostic sensitivity for penicilliosis.