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Article: Immunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosis

TitleImmunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosis
Authors
Issue Date2011
PublisherPublic 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?
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.
Persistent Identifierhttp://hdl.handle.net/10722/157663
ISSN
2013 Impact Factor: 3.534
2013 SCImago Journal Rankings: 1.724
PubMed Central ID
ISI Accession Number ID
Funding AgencyGrant Number
Ted Sun Foundation, Hong Kong
Research and Development Project of Guang Dong Province2010B090400498
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.

References

 

DC FieldValueLanguage
dc.contributor.authorWang, YFen_US
dc.contributor.authorCai, JPen_US
dc.contributor.authorWang, YDen_US
dc.contributor.authorDong, Hen_US
dc.contributor.authorHao, Wen_US
dc.contributor.authorJiang, LXen_US
dc.contributor.authorLong, Jen_US
dc.contributor.authorChan, Cen_US
dc.contributor.authorWoo, PCYen_US
dc.contributor.authorLau, SKPen_US
dc.contributor.authorYuen, KYen_US
dc.contributor.authorChe, XYen_US
dc.date.accessioned2012-08-08T08:52:03Z-
dc.date.available2012-08-08T08:52:03Z-
dc.date.issued2011en_US
dc.identifier.citationPLoS One, 2011, v. 6 n. 12, article no. e28796en_US
dc.identifier.issn1932-6203en_US
dc.identifier.urihttp://hdl.handle.net/10722/157663-
dc.description.abstractBACKGROUND: 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.en_US
dc.languageengen_US
dc.publisherPublic Library of Science. The Journal's web site is located at http://www.plosone.org/home.actionen_US
dc.relation.ispartofPLoS Oneen_US
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.subject.meshFungal Proteins - analysis - genetics - immunology - metabolism-
dc.subject.meshImmunoassay - methods-
dc.subject.meshMembrane Glycoproteins - analysis - genetics - immunology - metabolism-
dc.subject.meshMycoses - blood - diagnosis - microbiology-
dc.subject.meshPenicillium - genetics - growth and development - pathogenicity-
dc.titleImmunoassays based on Penicillium marneffei Mp1p derived from Pichia pastoris expression system for diagnosis of penicilliosisen_US
dc.typeArticleen_US
dc.identifier.emailChan, C: cmchan@hku.hken_US
dc.identifier.emailWoo, PCY: pcywoo@hkucc.hku.hken_US
dc.identifier.emailLau, SKP: skplau@hkucc.hku.hken_US
dc.identifier.emailYuen, KY: kyyuen@hkucc.hku.hk-
dc.identifier.emailChe, XY: chexiaoyan@126.com-
dc.identifier.authorityWoo, PCY=rp00430en_US
dc.identifier.authorityLau, SKP=rp00486en_US
dc.identifier.authorityYuen, KY=rp00366en_US
dc.description.naturepublished_or_final_versionen_US
dc.identifier.doi10.1371/journal.pone.0028796en_US
dc.identifier.pmid22205971-
dc.identifier.pmcidPMC3244411-
dc.identifier.scopuseid_2-s2.0-83755186102en_US
dc.identifier.hkuros204308-
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-83755186102&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume6en_US
dc.identifier.issue12, article no. e28796en_US
dc.identifier.isiWOS:000299113600035-
dc.publisher.placeUnited Statesen_US
dc.identifier.scopusauthoridChe, XY=7005743182en_US
dc.identifier.scopusauthoridYuen, KY=36078079100en_US
dc.identifier.scopusauthoridLau, SKP=7401596211en_US
dc.identifier.scopusauthoridWoo, PCY=7201801340en_US
dc.identifier.scopusauthoridChan, C=7404814453en_US
dc.identifier.scopusauthoridLong, J=36797221900en_US
dc.identifier.scopusauthoridJiang, LX=54784693100en_US
dc.identifier.scopusauthoridHao, W=7101686587en_US
dc.identifier.scopusauthoridDong, H=35770452600en_US
dc.identifier.scopusauthoridWang, YD=54785441500en_US
dc.identifier.scopusauthoridCai, JP=35557916700en_US
dc.identifier.scopusauthoridWang, YF=54785319300en_US

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