Article: Immunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosis
| Title | Immunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosis |
|---|---|
| Authors | Wang, YF3 Cai, JP3 Wang, YD3 Dong, H1 Hao, W3 Jiang, LX3 Long, J3 Chan, C2 Woo, PCY2 Lau, SKP2 Yuen, KY2 Che, XY3 |
| Issue Date | 2011 |
| Publisher | Public Library of Science. The Journal's web site is located at http://www.plosone.org/home.action |
| Citation | PLoS One, 2011, v. 6 n. 12, article no. e28796 [How to Cite?] DOI: http://dx.doi.org/10.1371/journal.pone.0028796 |
| Abstract | BACKGROUND: Penicillium marneffei is a dimorphic fungus endemic in Southeast Asia. It can cause fatal penicilliosis in humans, particularly in HIV-infected people. Diagnosis of this infection is difficult because its clinical manifestations are not distinctive. Specialized laboratory tests are necessary to establish a definitive diagnosis for successful management. We have demonstrated previously that a cell wall mannoprotein Mp1p, abundant in P. marneffei, is a potential biomarker for diagnosis of P. marneffei infections. In the present study, we describe immunoassays based on Mp1p derived from the yeast Pichia pastoris expression system. METHODOLOGY/PRINCIPAL FINDINGS: We generated monoclonal antibodies (MAbs) and rabbit polyclonal antibodies (PAbs) against Mp1p expressed in P. pastoris. Subsequently, we developed two Mp1p antigen capture ELISAs which employed MAbs for both the capture and detecting antibodies (MAb-MAb pair) or PAbs and MAbs as the capture and detecting antibodies (PAbs-MAb pair) respectively. The two Mp1p antigen ELISAs detected Mp1p specifically in cultures of P. marneffei yeast phase at 37-40 degrees C and had no cross-reaction with other tested pathogenic fungi. The sensitivities and specificities of the two antigen assays were found to be 55% (11/20) and 99.6% (538/540) for MAb-MAb Mp1p ELISA, and 75% (15/20) and 99.4% (537/540) for PAbs-MAb Mp1p ELISA performed using 20 sera with culture-confirmed penicilliosis, and 540 control sera from 15 other mycosis patients and 525 healthy donors. Meanwhile, we also developed an anti-Mp1p IgG antibody ELISA with an evaluated sensitivity of 30% (6/20) and a specificity of 98.5% (532/540) using the same sera. Furthermore, combining the results of Mp1p antigen and antibody detection improved the sensitivity of diagnosis to 100% (20/20). CONCLUSIONS/SIGNIFICANCE: Simultaneous detection of antigen and antibody using the immunoassays based on Mp1p derived from P. pastoris greatly improves detection sensitivity. The procedures should be useful for the routine diagnosis of penicilliosis. |
| ISSN | 1932-6203 2011 Impact Factor: 4.092 2011 SCImago Journal Rankings: 0.519 |
| DOI | http://dx.doi.org/10.1371/journal.pone.0028796 |
| PubMed Central ID | PMC3244411 |
| References | References in Scopus |
| dc.contributor.author | Wang, YF | ||||||
|---|---|---|---|---|---|---|---|
| dc.contributor.author | Cai, JP | ||||||
| dc.contributor.author | Wang, YD | ||||||
| dc.contributor.author | Dong, H | ||||||
| dc.contributor.author | Hao, W | ||||||
| dc.contributor.author | Jiang, LX | ||||||
| dc.contributor.author | Long, J | ||||||
| dc.contributor.author | Chan, C | ||||||
| dc.contributor.author | Woo, PCY | ||||||
| dc.contributor.author | Lau, SKP | ||||||
| dc.contributor.author | Yuen, KY | ||||||
| dc.contributor.author | Che, XY | ||||||
| dc.date.accessioned | 2012-08-08T08:52:03Z | ||||||
| dc.date.available | 2012-08-08T08:52:03Z | ||||||
| dc.date.issued | 2011 | ||||||
| dc.description.abstract | BACKGROUND: Penicillium marneffei is a dimorphic fungus endemic in Southeast Asia. It can cause fatal penicilliosis in humans, particularly in HIV-infected people. Diagnosis of this infection is difficult because its clinical manifestations are not distinctive. Specialized laboratory tests are necessary to establish a definitive diagnosis for successful management. We have demonstrated previously that a cell wall mannoprotein Mp1p, abundant in P. marneffei, is a potential biomarker for diagnosis of P. marneffei infections. In the present study, we describe immunoassays based on Mp1p derived from the yeast Pichia pastoris expression system. METHODOLOGY/PRINCIPAL FINDINGS: We generated monoclonal antibodies (MAbs) and rabbit polyclonal antibodies (PAbs) against Mp1p expressed in P. pastoris. Subsequently, we developed two Mp1p antigen capture ELISAs which employed MAbs for both the capture and detecting antibodies (MAb-MAb pair) or PAbs and MAbs as the capture and detecting antibodies (PAbs-MAb pair) respectively. The two Mp1p antigen ELISAs detected Mp1p specifically in cultures of P. marneffei yeast phase at 37-40 degrees C and had no cross-reaction with other tested pathogenic fungi. The sensitivities and specificities of the two antigen assays were found to be 55% (11/20) and 99.6% (538/540) for MAb-MAb Mp1p ELISA, and 75% (15/20) and 99.4% (537/540) for PAbs-MAb Mp1p ELISA performed using 20 sera with culture-confirmed penicilliosis, and 540 control sera from 15 other mycosis patients and 525 healthy donors. Meanwhile, we also developed an anti-Mp1p IgG antibody ELISA with an evaluated sensitivity of 30% (6/20) and a specificity of 98.5% (532/540) using the same sera. Furthermore, combining the results of Mp1p antigen and antibody detection improved the sensitivity of diagnosis to 100% (20/20). CONCLUSIONS/SIGNIFICANCE: Simultaneous detection of antigen and antibody using the immunoassays based on Mp1p derived from P. pastoris greatly improves detection sensitivity. The procedures should be useful for the routine diagnosis of penicilliosis. | ||||||
| dc.description.nature | published_or_final_version | ||||||
| dc.identifier.citation | PLoS One, 2011, v. 6 n. 12, article no. e28796 [How to Cite?] DOI: http://dx.doi.org/10.1371/journal.pone.0028796 | ||||||
| dc.identifier.doi | http://dx.doi.org/10.1371/journal.pone.0028796 | ||||||
| dc.identifier.hkuros | 204308 | ||||||
| dc.identifier.isi | WOS:000299113600035
Funding Information: This work was supported by the Ted Sun Foundation, Hong Kong and the Research and Development Project of Guang Dong Province (No. 2010B090400498). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. | ||||||
| dc.identifier.issn | 1932-6203 2011 Impact Factor: 4.092 2011 SCImago Journal Rankings: 0.519 | ||||||
| dc.identifier.issue | 12, article no. e28796 | ||||||
| dc.identifier.pmcid | PMC3244411 | ||||||
| dc.identifier.pmid | 22205971 | ||||||
| dc.identifier.scopus | eid_2-s2.0-83755186102 | ||||||
| dc.identifier.uri | http://hdl.handle.net/10722/157663 | ||||||
| dc.identifier.volume | 6 | ||||||
| dc.language | eng | ||||||
| dc.publisher | Public Library of Science. The Journal's web site is located at http://www.plosone.org/home.action | ||||||
| dc.publisher.place | United States | ||||||
| dc.relation.ispartof | PLoS One | ||||||
| dc.relation.references | References in Scopus | ||||||
| dc.rights | Creative Commons: Attribution 3.0 Hong Kong License | ||||||
| dc.subject.mesh | Fungal Proteins - analysis - genetics - immunology - metabolism | ||||||
| dc.subject.mesh | Immunoassay - methods | ||||||
| dc.subject.mesh | Membrane Glycoproteins - analysis - genetics - immunology - metabolism | ||||||
| dc.subject.mesh | Mycoses - blood - diagnosis - microbiology | ||||||
| dc.subject.mesh | Penicillium - genetics - growth and development - pathogenicity | ||||||
| dc.title | Immunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosis | ||||||
| dc.type | Article |
Author Affiliations
- Chinese National Human Genome Center at Shanghai
- The University of Hong Kong
- Zhujiang Hospital