About: In-plate recapturing of a dual-tagged recombinant Fasciola antigen (FhLAP) by a monoclonal antibody (US9) prevents non-specific binding in ELISA

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About: In-plate recapturing of a dual-tagged recombinant Fasciola antigen (FhLAP) by a monoclonal antibody (US9) prevents non-specific binding in ELISA Goto Sponge NotDistinct Permalink

An Entity of Type : schema:ScholarlyArticle, within Data Space : covidontheweb.inria.fr associated with source document(s)

Attributes	Values
type	Academic Article research paper schema:ScholarlyArticle
isDefinedBy	Covid-on-the-Web dataset
has title	In-plate recapturing of a dual-tagged recombinant Fasciola antigen (FhLAP) by a monoclonal antibody (US9) prevents non-specific binding in ELISA
Creator	Mezo, Mercedes Paniagua, Esperanza Romarís, Fernanda Orbegozo-Medina, Ricardo Perteguer, María Gonzá Lez-Warleta, Marta Gá Rate, Teresa Herná Ndez-Gonzá Lez, Ana Martínez-Serná Ndez, Victoria Ubeira Id, Florencio
Source	Medline; PMC
abstract	Recombinant proteins expressed in E. coli are frequently purified by immobilized metal affinity chromatography (IMAC). By means of this technique, tagged proteins containing a polyhistidine sequence can be obtained up to 95% pure in a single step, but some host proteins also bind with great affinity to metal ions and contaminate the sample. A way to overcome this problem is to include a second tag that is recognized by a preexistent monoclonal antibody (mAb) in the gene encoding the target protein, allowing further purification. With this strategy, the recombinant protein can be directly used as target in capture ELISA using plates sensitized with the corresponding mAb. As a proof of concept, in this study we engineered a Trichinella-derived tag (MTFSVPIS, recognized by mAb US9) into a His-tagged recombinant Fasciola antigen (rFhLAP) to make a new chimeric recombinant protein (rUS9-FhLAP), and tested its specificity in capture and indirect ELISAs with sera from sheep and cattle. FhLAP was selected since it was previously reported to be immunogenic in ruminants and is expressed in soluble form in E. coli, which anticipates a higher contamination by host proteins than proteins expressed in inclusion bodies. Our results showed that a large number of sera from non-infected ruminants (mainly cattle) reacted in indirect ELISA with rUS9-FhLAP after single-step purification by IMAC, but that this reactivity disappeared testing the same antigen in capture ELISA with mAb US9. These results demonstrate that the 6XHis and US9 tags can be combined when double purification of recombinant proteins is required.
has issue date	2019-02-01(xsd:dateTime)
bibo:doi	10.1371/journal.pone.0211035
bibo:pmid	30707711
has license	cc-by
sha1sum (hex)	7b5d559562d07c48dfbba720712d27a53ee81504
schema:url	https://doi.org/10.1371/journal.pone.0211035
resource representing a document's title	In-plate recapturing of a dual-tagged recombinant Fasciola antigen (FhLAP) by a monoclonal antibody (US9) prevents non-specific binding in ELISA
has PubMed Central identifier	PMC6358068
has PubMed identifier	30707711
schema:publication	PLoS One
resource representing a document's body	covid:7b5d559562d07c48dfbba720712d27a53ee81504#body_text
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