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Micro-bead injection spectroscopy for label-free automated determination of immunoglobulin G in human serum

2017Journal article Open access
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dc.contributor.authorRamos, Inês I.
dc.contributor.authorMagalhães, Luís M.
dc.contributor.authorBarreiros, Luisa
dc.contributor.authorReis, Salette
dc.contributor.authorLima, José L. F. C.
dc.contributor.authorSegundo, Marcela A.
dc.date.accessioned2019-07-03T16:32:50Z
dc.date.available2019-07-03T16:32:50Z
dc.date.issued2017
dc.description.abstractImmunoglobulin G (IgG) represents the major fraction of antibodies in healthy adult human serum, and deviations from physiological levels are a generic marker of disease corresponding to different pathologies. Therefore, screening methods for IgG evaluation are a valuable aid to diagnostics. The present work proposes a rapid, automatic, and miniaturized method based on UV-vis micro-bead injection spectroscopy (μ-BIS) for the real-time determination of human serum IgG with label-free detection. Relying on attachment of IgG in rec-protein G immobilized in Sepharose 4B, a bioaffinity column is automatically assembled, where IgG is selectively retained and determined by on-column optical density measurement. A "dilution-and-shoot" approach (50 to 200 times) was implemented without further sample treatment because interferences were flushed out of the column upon sample loading, with minimization of carryover and cross-contamination by automatically discarding the sorbent (0.2 mg) after each determination. No interference from human serum albumin at 60 mg mL-1 in undiluted sample was found. The method allowed IgG determination in the range 100-300 μg mL-1 (corresponding to 5.0-60 mg mL-1 in undiluted samples), with a detection limit of 33 μg mL-1 (1.7 mg mL-1 for samples, dilution factor of 50). RSD values were < 9.4 and < 11.7%, for intra and inter-assay precision, respectively, while recovery values for human serum spiked with IgG at high pathological levels were 97.8-101.4%. Comparison to commercial ELISA kit showed no significant difference for tested samples (n = 8). Moreover, time-to-result decreased from several hours to < 5 min and analysis cost decreased 10 times, showing the potential of the proposed approach as a point-of-care method. Graphical abstract Micro-Bead Injection Spectroscopy method for real time, automated and label-free determination of total serum human Immunoglobulin G (IgG). The method was designed for Lab-on-Valve (LOV) platforms using a miniaturised protein G bioaffinity separative approach. IgG are separated from serum matrix components upon quantification with low non-specific binding in less than 5 min.pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.doi10.1007/s00216-017-0601-6pt_PT
dc.identifier.urihttp://hdl.handle.net/10400.22/14242
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherSpringerpt_PT
dc.relationPOCI/01/0145/FEDER/007728pt_PT
dc.relationPOCI/01/0145/FEDER/007265pt_PT
dc.relation.publisherversionhttps://link.springer.com/article/10.1007%2Fs00216-017-0601-6pt_PT
dc.subjectBacterial Proteinspt_PT
dc.subjectEquipment Designpt_PT
dc.subjectHumanspt_PT
dc.subjectImmobilized Proteinspt_PT
dc.subjectImmunoglobulin Gpt_PT
dc.subjectSepharosept_PT
dc.subjectSpectrum Analysispt_PT
dc.titleMicro-bead injection spectroscopy for label-free automated determination of immunoglobulin G in human serumpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/5876/UID%2FMulti%2F04378%2F2013/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/5876/UID%2FQUI%2F50006%2F2013/PT
oaire.citation.endPage988pt_PT
oaire.citation.issue3pt_PT
oaire.citation.startPage981pt_PT
oaire.citation.titleAnalytical and Bioanalytical Chemistrypt_PT
oaire.citation.volume410pt_PT
oaire.fundingStream5876
oaire.fundingStream5876
person.familyNameBarreiros
person.givenNameLuisa
person.identifier.ciencia-id611F-E0C5-0230
person.identifier.orcid0000-0003-3481-5809
person.identifier.ridD-7950-2013
person.identifier.scopus-author-id6508205485
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT
relation.isAuthorOfPublication1e66bacc-64de-4ecb-96b7-4c0e366cba57
relation.isAuthorOfPublication.latestForDiscovery1e66bacc-64de-4ecb-96b7-4c0e366cba57
relation.isProjectOfPublicationa1891cb0-b820-4fd0-bc6c-e8918d4d4ce6
relation.isProjectOfPublication5c06a623-3499-408c-aef9-0a1f68cb1d02
relation.isProjectOfPublication.latestForDiscoverya1891cb0-b820-4fd0-bc6c-e8918d4d4ce6

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