Evidence map›Paper›PMID 38785994›Full record

ArticleBiomolecules2024

A Practical Guide for the Quality Evaluation of Fluobodies/Chromobodies.

Urša Štrancar, Claudia D'Ercole, Lucia Cikatricisová, Mirna Nakić, Matteo De March, Ario de Marco

Abstract read
In one paragraph

Article in Biomolecules, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

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1 · What the graph read from it

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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.

2 · The registry

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3 · Its place in the literature

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0 citing papers in PubMed.

No citing paper in PubMed yet.

4 · The record

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PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.

5 · Who and what money

Authors and funding

6 authors.

Urša ŠtrancarLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.ORCID 0009-0009-2266-3052
Claudia D'ErcoleLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.ORCID 0009-0008-0281-8697
Lucia CikatricisováLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.ORCID 0009-0002-2997-5576
Mirna NakićLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.
Matteo De MarchLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.ORCID 0000-0002-3447-957X
Ario de MarcoLaboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska cesta 13, Rožna Dolina, 5000 Nova Gorica, Slovenia.ORCID 0000-0001-7729-819X

Funding

Javne agencije za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije P3-0428, N4-0282, N4-0325, J4-50144
6 · The paper itself

Abstract

backgroundFluorescent proteins (FPs) are pivotal reagents for flow cytometry analysis or fluorescent microscopy. A new generation of immunoreagents (fluobodies/chromobodies) has been developed by fusing recombinant nanobodies to FPs.

methodsWe analyzed the quality of such biomolecules by a combination of gel filtration and SDS-PAGE to identify artefacts due to aggregation or material degradation.

resultsIn the SDS-PAGE run, unexpected bands corresponding to separate fluobodies were evidenced and characterized as either degradation products or artefacts that systematically resulted in the presence of specific FPs and some experimental conditions. The elimination of N-terminal methionine from FPs did not impair the appearance of FP fragments, whereas the stability and migration characteristics of some FP constructs were strongly affected by heating in loading buffer, which is a step samples undergo before electrophoretic separation.

conclusionsIn this work, we provide explanations for some odd results observed during the quality control of fluobodies and summarize practical suggestions for the choice of the most convenient FPs to fuse to antibody fragments.

Indexed as

Electrophoresis, Polyacrylamide GelChromatography, GelFlow CytometryHumansQuality ControlSingle-Domain AntibodiesSingle-Domain Antibodiesalternative start codonsfluobodiesfluorescent proteinsprotein degradation

Identifiers

PMID38785994
PMCPMC11117837

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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.