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Statements

Subject Item
n2:RIV%2F61989592%3A15310%2F14%3A33151657%21RIV15-MSM-15310___
rdf:type
n4:Vysledek skos:Concept
rdfs:seeAlso
http://www.sciencedirect.com/science/article/pii/S0021967314017993
dcterms:description
Apidaecins represent an important group of antimicrobial peptides occurring in honey bee hemolymph, where they play an important role as key components of humoral immunity. The present study demonstrates the development of a highly sensitive assay for apidaecin 1 isoforms quantification in the hemolymph or body parts from honey bee individuals. The analytical protocol comprises apidaecins 1 purification and enrichment steps by weak cation-exchange chromatography (WCX) in laboratory-made WCX-Tip microcolumns combined with a desalting step on a reversed-phase sorbent (C8) carried in StageTips. Apidaecin-enriched fraction was analyzed by a reversed-phase based nanoliquid chromatography (C4) separation coupled with high-resolution mass spectrometry. The method performance was validated in its specificity, linearity (0-5 pmol), recovery (?45%), precision ({10% at 0.1 pmol), limit of detection (?50 fmol), limit of quantification (0.1 pmol) and sample stability. The method was successfully applied to analyze the content of apidaecin 1 isoforms in the following samples: hemolymph - 13.0 ng/mýL (95% confidence interval of 7.5-18.6 ng/mýL), thoraxes - 36.2 ng/unit (95% CI of 18.9-53.6 ng/unit) and heads - 12.9 ng/unit (95% CI of 9.1-16.7 ng/unit). Freshly emerged bees had apidaecin 1 isoforms levels below the limit of detection. Thus it was possible to use them as a competitive matrix for calibration standards to prevent losses of highly basic apidaecins. This new protocol for apidaecin 1 isoforms quantification represents a promising tool to study the role of apidaecins in honey bee immunity and can be considered as a proof-of-concept for the development of sensitive quantification methods for basic antimicrobial peptides in various organisms. Apidaecins represent an important group of antimicrobial peptides occurring in honey bee hemolymph, where they play an important role as key components of humoral immunity. The present study demonstrates the development of a highly sensitive assay for apidaecin 1 isoforms quantification in the hemolymph or body parts from honey bee individuals. The analytical protocol comprises apidaecins 1 purification and enrichment steps by weak cation-exchange chromatography (WCX) in laboratory-made WCX-Tip microcolumns combined with a desalting step on a reversed-phase sorbent (C8) carried in StageTips. Apidaecin-enriched fraction was analyzed by a reversed-phase based nanoliquid chromatography (C4) separation coupled with high-resolution mass spectrometry. The method performance was validated in its specificity, linearity (0-5 pmol), recovery (?45%), precision ({10% at 0.1 pmol), limit of detection (?50 fmol), limit of quantification (0.1 pmol) and sample stability. The method was successfully applied to analyze the content of apidaecin 1 isoforms in the following samples: hemolymph - 13.0 ng/mýL (95% confidence interval of 7.5-18.6 ng/mýL), thoraxes - 36.2 ng/unit (95% CI of 18.9-53.6 ng/unit) and heads - 12.9 ng/unit (95% CI of 9.1-16.7 ng/unit). Freshly emerged bees had apidaecin 1 isoforms levels below the limit of detection. Thus it was possible to use them as a competitive matrix for calibration standards to prevent losses of highly basic apidaecins. This new protocol for apidaecin 1 isoforms quantification represents a promising tool to study the role of apidaecins in honey bee immunity and can be considered as a proof-of-concept for the development of sensitive quantification methods for basic antimicrobial peptides in various organisms.
dcterms:title
A sensitive quantification of the peptide apidaecin 1 isoforms in single bee tissues using a weak cation exchange pre-separation and nanocapillary liquid chromatography coupled with mass spectrometry A sensitive quantification of the peptide apidaecin 1 isoforms in single bee tissues using a weak cation exchange pre-separation and nanocapillary liquid chromatography coupled with mass spectrometry
skos:prefLabel
A sensitive quantification of the peptide apidaecin 1 isoforms in single bee tissues using a weak cation exchange pre-separation and nanocapillary liquid chromatography coupled with mass spectrometry A sensitive quantification of the peptide apidaecin 1 isoforms in single bee tissues using a weak cation exchange pre-separation and nanocapillary liquid chromatography coupled with mass spectrometry
skos:notation
RIV/61989592:15310/14:33151657!RIV15-MSM-15310___
n3:aktivita
n16:P
n3:aktivity
P(LO1204)
n3:cisloPeriodika
DEC
n3:dodaniDat
n12:2015
n3:domaciTvurceVysledku
n6:9896821 n6:5665442 n6:9708790 n6:5208564
n3:druhVysledku
n18:J
n3:duvernostUdaju
n19:S
n3:entitaPredkladatele
n11:predkladatel
n3:idSjednocenehoVysledku
1159
n3:idVysledku
RIV/61989592:15310/14:33151657
n3:jazykVysledku
n7:eng
n3:klicovaSlova
Quantification; Mass spectrometry; Nanoflow liquid chromatography; Honey bee; Apidaecin; Antimicrobial peptide
n3:klicoveSlovo
n9:Antimicrobial%20peptide n9:Honey%20bee n9:Quantification n9:Mass%20spectrometry n9:Apidaecin n9:Nanoflow%20liquid%20chromatography
n3:kodStatuVydavatele
NL - Nizozemsko
n3:kontrolniKodProRIV
[3669AA2159A1]
n3:nazevZdroje
Journal of Chromatography A
n3:obor
n10:CE
n3:pocetDomacichTvurcuVysledku
4
n3:pocetTvurcuVysledku
4
n3:projekt
n5:LO1204
n3:rokUplatneniVysledku
n12:2014
n3:svazekPeriodika
1374
n3:tvurceVysledku
Šebela, Marek Danihlík, Jiří Petřivalský, Marek Lenobel, René
n3:wos
000347605000015
s:issn
0021-9673
s:numberOfPages
11
n20:doi
10.1016/j.chroma.2014.11.041
n17:organizacniJednotka
15310