Wang Yanfang, Zeng Lei, Chen Xuedong, Hao Wei, Yang Mei, Cai Jianpiao, Wang Yadi, Yuan Guoyong, Che Xiaoyan
Center for Clinical Laboratory, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2013 Mar;33(3):439-43.
To establish an immunological method for detecting antibodies of Penicillium marneffei.
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.
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).
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.
建立检测马尔尼菲青霉抗体的免疫学方法。
马尔尼菲青霉重组Mp1p蛋白在毕赤酵母中表达，并用改良过碘酸钠法标记辣根过氧化物酶（Mp1p-HRP）。通过确定Mp1p蛋白的最佳包被浓度和检测蛋白Mp1p-HRP的浓度，建立双抗原夹心酶联免疫吸附测定（ELISA）法。通过检测100份健康供者血清样本、15份培养确诊的马尔尼菲青霉病患者样本和21份培养确诊的其他真菌感染患者样本中的Mp1p抗体，评估该检测方法的敏感性和特异性。
成功建立了检测Mp1p特异性抗体的双抗原夹心ELISA法。该检测方法检测健康供者和其他真菌感染患者血清中Mp1p特异性抗体的特异性为100%（121/121）。15份培养确诊的马尔尼菲青霉病患者血清样本检测结果显示，2份样本Mp1p抗体阳性，12份样本Mp1p抗原阳性；Mp1p抗原和抗体检测结果相结合，明显提高诊断敏感性至93.3%（14/15）。
双抗原夹心ELISA法检测Mp1p特异性抗体具有较高特异性，同时检测Mp1p抗原和抗体可提高马尔尼菲青霉病的诊断敏感性。
