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Three ml of distilled water was added on the polymeric film as release bulk.
Three mL of sodium chloride 0.9% were added, followed by 4 min of mechanical homogenization with a Stomacher 80 Biomaster (Seward, Worthing, UK).
Briefly, three ml of freshly obtained heparinzed human blood was gently mixed, poured into sterile 15 ml falcon tube and centrifuged for 5 min at 850 × g.
Three mL of broth collected at T0 and at the end of the process were centrifuged and supernatants were ultrafiltered on 3 kDa centricon devices (Millipore, Bedford, MA, USA) at 5000×g and the flow through was used for analyses.
Three ml of cultures of the indicated strains were harvested at OD600 = 0.8.
Three ml (10 MOI) of this supernatant was added to SH-SY5Y cells.
Three ml of peritoneal fluid were obtained from each mouse at 24 hours post-CLP.
Three mL were then taken and treated with 6 mL of RNAprotect Bacteria Reagent (Qiagen, Hilden, Germany).
Three ml of 5 M sodium perchlorate was added and incubated for 1 h at ambient temperature.
Three ml of blood was collected from each patient each in EDTA containing tube and in a plain tube.
Three ml fractions were collected and assayed in triplicate for cell-cycle inhibiting activity using the BrdU assay.
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