Human RNA Isolation from Tempus Tubes and quantitative Real-Time
PCR.
At Baseline visit whole blood was collected in Tempus® Blood RNA Tubes
(Life Technologies™, GmbH, Darmstadt, Germany) and RNA was extracted
with the MagMax for Stabilized Blood Tubes RNA Isolation Kit. Synthesis
of cDNA and consequently real time-PCR were performed as described for
murine cells below with the following primers andsequences: hHPRT
(5´-TGA CAC TGG CAA AAC AAT GCA-3´, 5´-GGT CCT TTT CAC CAG CAA GCT-3´),
PD-1 (fw: 5´-CAG TTC CAA ACC CTG GTG GT- 3´; rev: 5´-GGC TCC TAT TGT CCC
TCG TG- 3´).
FEV1 .
The Spirometry method for the lung function analysis was performed by
using a Vyaire Bodyplethysmograph (Chicago,USA). ERS Criteria were
followed during the study. The quality control was performed by an
experienced technician and a physician and supported by the
software.FEV1 (=Forced expiratory volume in 1 second) and FVC (=Forced
vital capacity) were measured at Baseline visit (B0) by using
spirometry. After a period of normal breathing the participant should
inhale maximal, directly followed by maximal and fast exhalation. The
volume exhaled in one second is FEV1. The total exhaled volume is FVC.
The ratio FVC/FEV1 is stated as FEV1%.