Nasal Respiratory Epithelial Cell Collection and RNA
Isolation
Nasal brushings were obtained from the mucosal surface using a 0.8 mm
interdental brush (TePe, Ireland). The nasal brush was then transferred
immediately to 600 µl lysis buffer (Exiqon, Germany) containing 6 µl
β-mercaptoethanol (Sigma-Aldrich, USA) and gently pushed through a
sterile pipette tip to release the lysate into the lysis buffer.
Brushings from two nostrils were pooled. Total RNA was isolated usingmiRCURYTM RNA Isolation Kit (Exiqon, Germany).
Lysate transferred to the lysis buffer was homogenized and after
Proteinase K (Thermo Fisher Scientific, USA) treatment, it was
transferred to mini spin column (Exiqon, Germany). After DNase I
treatment (Bio-Rad), total RNA was eluted in RNase free
H2O. Total RNA was stored at -80 0C.
RNA quality and quantity was determined using NanoDrop ND-1000 (Thermo
Fisher Scientific, USA).