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2017 ; 65
(3
): 706-712
Nephropedia Template TP
gab.com Text
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Twit Text #
English Wikipedia
Validation of an enzyme-linked immunosorbent assay for the quantification of
citrullinated histone H3 as a marker for neutrophil extracellular traps in human
plasma
#MMPMID28161762
Thålin C
; Daleskog M
; Göransson SP
; Schatzberg D
; Lasselin J
; Laska AC
; Kallner A
; Helleday T
; Wallén H
; Demers M
Immunol Res
2017[Jun]; 65
(3
): 706-712
PMID28161762
show ga
There is an emerging interest in the diverse functions of neutrophil
extracellular traps (NETs) in a variety of disease settings. However, data on
circulating NETs rely largely upon surrogate NET markers such as cell-free DNA,
nucleosomes, and NET-associated enzymes. Citrullination of histone H3 by peptidyl
arginine deiminase 4 (PAD4) is central for NET formation, and citrullinated
histone H3 (H3Cit) is considered a NET-specific biomarker. We therefore aimed to
optimize and validate a new enzyme-linked immunosorbent assay (ELISA) to quantify
the levels of H3Cit in human plasma. A standard curve made of in vitro
PAD4-citrullinated histones H3 allows for the quantification of H3Cit in plasma
using an anti-histone antibody as capture antibody and an anti-histone H3
citrulline antibody for detection. The assay was evaluated for linearity,
stability, specificity, and precision on plasma samples obtained from a human
model of inflammation before and after lipopolysaccharide injection. The results
revealed linearity and high specificity demonstrated by the inability of
detecting non-citrullinated histone H3. Coefficients of variation for intra- and
inter-assay variability ranged from 2.1 to 5.1% and from 5.8 to 13.5%,
respectively, allowing for a high precision. Furthermore, our results support an
inflammatory induction of a systemic NET burden by showing, for the first time,
clear intra-individual elevations of plasma H3Cit in a human model of
lipopolysaccharide-induced inflammation. Taken together, our work demonstrates
the development of a new method for the quantification of H3Cit by ELISA that can
reliably be used for the detection of NETs in human plasma.