Antithrombin III as predictive indicator of survival
BAL samples were filtered through sterile gauze to remove mucus. Cell count was performed by cytocentrifuge smear (600 rpm for 5 min) using May-Grunewald Giemsa staining (DiaPath, Italy).
Cell viability was determined by Trypan blue exclusion. Flow cytometric analysis was performed using monoclonal antibodies (BD Multitest™ 6-color TBNK, San Jose, CA, USA), including CD3 FITC-labelled, CD16/CD56 PE-labelled, CD45 PerCP-Cy5.5-labelled, CD4 PE-Cy7-labelled, CD19 APC-labelled and CD8 APC-Cy7-labelled. About 1x106 cells were incubated with antibodies for 30 minutes, and then washed with 500 μl RPMI-1640 medium.