P2XR mRNA is expressed in human CF and non-CF airway epithelial cell lines and in primary cultures. (a) P2XR PCR products from 16HBE14o^– non-CF human bronchial epithelial cells, CFBE41o^– CF human bronchial epithelial cells, ΣCFTE-29o^– CF human tracheal epithelial cells (shown in b), a CFNP epithelial primary culture, Calu-3 non-CF human submucosal gland serous cells, and human umbilical vein endothelial cells (HUVEC). NC, no cDNA (control). (b) Comparison of positive airway epithelial cell reactions to T84 non-CF human colon carcinoma cells and PANC-1 non-CF human pancreatic epithelial cells. (c) Positive reactions from the 16HBE14o^– sample compared with CFPAC-1 CF human pancreatic epithelial cells, IMCD renal inner medullary collecting-duct cells, and HUVEC. (d) Positive reactions for A549 non-CF human alveolar lung cells and Beas2B non-CF human bronchial epithelial cells vs. control (No RT). Positive reactions for CFPAC-1 and 16HBE14o^– cells are also shown as positive controls for comparison. Beas2B 1 and Beas2B 2 are two different cDNA samples from the same RNA sample. (e) PCR controls on 16HBE14o^– total RNA sample. No P2XR PCR product was derived from amplification of total RNA (–/–) or RNase-free, DNase-treated total RNA (DNase), whereas positive reactions occurred in total RNA reverse-transcribed to cDNA with (DNase/RT) and without (RT) DNase treatment. All amplifications of epithelial cell samples were performed 2–3 times. Note: Analysis of the sites that the degenerate P2X PCR primers recognized within the genomic sequence of the mouse P2X3 gene (28) indicated that our primers spanned at least 2 introns; amplification of genomic DNA would produce a PCR product at least 2 kb larger than the 330-bp expected size. Moreover, in the P2X3 genomic sequence, our reverse primer spanned an exon/intron boundary (intron ∼25 kb in size), rendering amplification of genomic DNA improbable.