Barrera Coralie, Millon Laurence, Rognon Bénédicte, Quadroni Manfredo, Roussel Sandrine, Dalphin Jean-Charles, Court-Fortune Isabelle, Caillaud Denis, Jouneau Stephane, Fellrath Jean-Marc, Zaugg Christophe, Reboux Gabriel, Monod Michel
UMR6249 Chrono-environnement, University of Franche-Comté, Besançon, France; Parasitology-Mycology Department, University Hospital of Besançon, Besançon, France.
Proteomics Clin Appl. 2014 Dec;8(11-12):971-81. doi: 10.1002/prca.201400024. Epub 2014 Nov 4.
Saccharopolyspora rectivirgula is the principal cause of farmer's lung disease (FLD). Serodiagnosis is based on immunoprecipitation techniques or enzyme immunoassays with homemade crude antigens and is not standardized. We aimed to produce specific recombinant antigens for the development of a standardized ELISA.
We recruited 41 patients and 43 healthy exposed controls from five university hospital pneumology departments in France and Switzerland. S. rectivirgula proteins were extracted, separated by 2D electrophoresis, and subjected to Western blotting, with sera from FLD patients or controls. FLD-specific proteins were identified by MS and were produced as recombinant antigens. The diagnostic performance of ELISA tests using the recombinant antigens was assessed with all the sera from FLD patients and controls.
We identified 25 FLD-specific proteins, some of which play important roles in transport, nutrition, or virulence. We produced 17 of these proteins as recombinant antigens and assessed their suitability for inclusion in the ELISA test. A combination of three of these proteins (SR1FA, SR17, and SR22) proved remarkably effective at discriminating between patients and controls, with a sensitivity of 83% and a specificity of 77%.
The recombinant antigens produced in this study constitute a major step toward the improvement of diagnostic performance and the standardization of FLD serodiagnosis.
直丝链孢菌是农民肺疾病(FLD)的主要病因。血清诊断基于免疫沉淀技术或使用自制粗抗原的酶免疫测定法,且未标准化。我们旨在生产特异性重组抗原来开发标准化的酶联免疫吸附测定(ELISA)。
我们从法国和瑞士的五个大学医院呼吸科招募了41例患者和43名健康暴露对照者。提取直丝链孢菌蛋白,通过二维电泳分离,并用FLD患者或对照者的血清进行蛋白质印迹分析。通过质谱鉴定FLD特异性蛋白,并将其作为重组抗原生产。使用重组抗原的ELISA检测的诊断性能用FLD患者和对照者的所有血清进行评估。
我们鉴定出25种FLD特异性蛋白,其中一些在运输、营养或毒力方面起重要作用。我们将其中17种蛋白作为重组抗原生产,并评估它们是否适合用于ELISA检测。其中三种蛋白(SR1FA、SR17和SR22)的组合在区分患者和对照方面证明非常有效,灵敏度为83%,特异性为77%。
本研究中产生的重组抗原是朝着提高诊断性能和FLD血清诊断标准化迈出的重要一步。