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Chromatographic behaviour of monoclonal antibodies against wild-type amidase from Pseudomonasaeruginosa on immobilized metal chelates

2011-12Journal article Open access
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dc.contributor.authorMartins, Sónia
dc.contributor.authorKarmali, Amin
dc.contributor.authorSerralheiro, Maria Luísa
dc.date.accessioned2013-01-31T12:22:50Z
dc.date.available2013-01-31T12:22:50Z
dc.date.issued2011-12
dc.description.abstractThe aim of this work was to devise a one-step purification procedure for monoclonal antibodies (MAbs) of IgG class by immobilized metal affinity chromatography (IMAC). Therefore, several stationary phases were prepared containing immobilized metal chelates in order to study the chromatographic behaviour of MAbs against wild-type amidase from Pseudomonas aeruginosa. Such MAbs adsorbed to Cu(II), Ni(II), Zn(II) and Co(II)-IDA agarose columns. The increase in ligand concentration and the use of longer spacer arms and higher pH values resulted in higher adsorption of MAbs into immobilized metal chelates. The dynamic binding capacity and the maximum binding capacity were 1.33 +/- 0.015 and 3.214 +/- 0.021 mg IgG/mL of sedimented commercial matrix, respectively. A K(D) of 4.53 x 10(-7) M was obtained from batch isotherm measurements. The combination of tailor-made stationary phases of IMAC and the correct selection of adsorption conditions permitted a one-step purification procedure to be devised for MAbs of IgG class. Culture supernatants containing MAbs were purified by IMAC on commercial-Zn(II) and EPI-30-IDA-Zn(II) Sepharose 6B columns and by affinity chromatography on Protein A-Sepharose CL-4B. This MAb preparation revealed on SDS-PAGE two protein bands with M(r) of 50 and 22 kDa corresponding to the heavy and light chains, respectively. Copyright (C) 2011 John Wiley & Sons, Ltd.por
dc.identifier.citationMARTINS, Sónia; KARMALI, Amin; SERRALHEIRO, Maria Luísa - Chromatographic behaviour of monoclonal antibodies against wild-type amidase from Pseudomonasaeruginosa on immobilized metal chelates. Biomedical Chromatograhy. ISSN 0269-3879. Vol. 25, n.º 12 (2011) p. 1327-1337.por
dc.identifier.issn0269-3879
dc.identifier.urihttp://hdl.handle.net/10400.21/2118
dc.language.isoengpor
dc.peerreviewedyespor
dc.publisherWiley-Blackwellpor
dc.subjectMonoclonal antibodies of IgG classpor
dc.subjectWild-type amidase from pseudomonas aeruginosapor
dc.subjectImmobilized metal affinity chromatographypor
dc.subjectZn(2+) Ionspor
dc.subjectEpichlorohydrinpor
dc.subjectIon affinity-chromatographypor
dc.subjectTomato pomacepor
dc.subjectPurificationpor
dc.subjectProteinspor
dc.subjectBindingpor
dc.subjectMutantpor
dc.titleChromatographic behaviour of monoclonal antibodies against wild-type amidase from Pseudomonasaeruginosa on immobilized metal chelatespor
dc.typejournal article
dspace.entity.typePublication
oaire.citation.conferencePlaceMaldenpor
oaire.citation.endPage1337por
oaire.citation.issue12por
oaire.citation.startPage1327por
oaire.citation.titleBiomedical Chromatograhypor
oaire.citation.volume25por
person.familyNameKarmali
person.givenNameAmin
person.identifier.orcid0000-0003-0419-401X
person.identifier.ridE-4787-2011
person.identifier.scopus-author-id6603778216
rcaap.rightsrestrictedAccesspor
rcaap.typearticlepor
relation.isAuthorOfPublication23975b4b-3234-4139-9675-89ea6f9a2332
relation.isAuthorOfPublication.latestForDiscovery23975b4b-3234-4139-9675-89ea6f9a2332

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