Computational protocol: Characterization of Microparticles after Hepatic Ischemia-Reperfusion Injury

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Protocol publication

[…] Whole blood was obtained by cardiac puncture using heparin as an anticoagulant. Plasma was labeled for MPs of specific cell origin as previously described . Forward and side-channels were set to analyze particles using 0.3, 1.1, and 3.0 µm latex beads for calibration (). The forward and side scatter gate was set based on size and vesicles within the 0.3–1.0 µm were identified as MPs () and further analyzed. MPs were then assessed with both source specific markers and Annexin V (BD Pharmingen, San Diego, California, USA) which stains for the presence of phosphatidylserine, a cell surface marker linked to apoptosis . Platelet-derived MP levels were assessed using the platelet-specific marker Anti-CD41 (clone: MWReg30, BD Pharmingen, San Diego, California, USA) (). Neutrophil-derived MP levels were assessed using neutrophil specific marker, Anti-Ly-6G (clone: 1A8, BD Pharmingen, San Diego, California, USA) (). Endothelial-derived MP levels were assessed using endothelial cell specific marker Anti-CD-62E (clone 10E9.6, BD Pharmingen, San Diego, California, USA) (). Flow cytometry data acquisition and analysis were performed using a Beckman Coulter Epic flow cytometer with Kaluza software (Indianapolis, IN, USA). [...] All data are expressed as mean ± standard error of the mean (SEM). Data were analyzed with a one-way analysis of variance with subsequent Student-Newman-Keuls test. Differences were considered significant when P<0.05. All statistical analysis was performed using SigmaPlot, Version 11 (Systat Software, Inc. San Jose, California, USA). […]

Pipeline specifications

Software tools Kaluza, SigmaPlot
Applications Miscellaneous, Flow cytometry
Organisms Mus musculus, Homo sapiens
Diseases Wounds and Injuries, Chemical and Drug Induced Liver Injury