Carcinoembryonic antigen (CEA)-related cell adhesion molecules are co-expressed in the human lung and their expression can be modulated in bronchial epithelial cells by non-typable Haemophilus influenzae, Moraxella catarrhalis, TLR3, and type I and II interferons

Klaile, Esther; Klassert, Tilman E; Scheffrahn, Inka; Müller, Mario M; Heinrich, Annina; Heyl, Kerstin A; Dienemann, Hendrik; Grünewald, Christiane; Bals, Robert; Singer, Bernhard B; Slevogt, Hortense

To quantify the relative expression of each gene, we used a CAS-1200 pipetting robot (Qiagen) to set up the qPCRreactions and a Corbett Rotor-Gene 6000 (Qiagen) as Real-Time qPCR apparatus. Each sample was analyzed in duplicate in a total reaction volume of 20 μl containing 10 μl of 2× SensiMix SYBR Master Mix (Bioline) and 0.2 μM of each primer. The cycling conditions included an initial step of 95°C for 10 min followed by 40 cycles of 95°C for 15 s, 60°C for 20 s and 72°C for 20 s. For each experiment, an RT-negative sample was included as a control. Melting curve analysis and size verification by electrophoresis was used to confirm the specificity of the qPCR reactions.