Fig. 3

Kinetics of NET production in vitro. The kinetics of cfDNA, as determined by qPCR, in the supernatant of control non-stimulated and stimulated by 1 mg/ml LPS or 100 nM PMA human neutrophils from a healthy donor following incubation for up to 5 h at 37 °C in cell culture: A total and long (> 320 bp) nuclear cf-nDNA concentration; B total and long (> 310 bp) mitochondrial cf-mtDNA concentration; C MNR (ratio of mitochondrial to nuclear DNA concentration). D Kinetics of NE and MPO in the supernatant of the same control non-stimulated and stimulated by 1 mg/ml LPS or 100 nM PMA. Data are represented as mean ± SD. E NET production: the cfDNA size profiles of the supernatant of human neutrophils before incubation and stimulation (PMA 0), and following stimulation by PMA 100 nM and incubation at 37 °C in cell culture for 2 h (PMA 2 h) and 5 h (PMA 5 h), as determined by sWGS. F Analysis of cfDNA size profile by capillary electrophoresis (Fragment Analyzer) in the supernatant of control non-stimulated (CTR 5 h) and stimulated by LPS (LPS 5 h) and PMA (PMA 5 h) neutrophils following 5 h incubation in cell culture. G, H The fluorescent microscopic image of Hoechst stained neutrophils stimulated by LPS (H) and control neutrophils (G) following 5 h incubation in cell culture. Examination of DNA content using Y chromosome analysis