Specimens were examined using a Nikon T1000U inverted phase-contrast microscope (Nikon UK Ltd; Surrey, England) and a × 40 objective lens. Only undisrupted ciliated strips of > 50 μm in length were studied. Beating ciliated edges were recorded using a digital video camera (Troubleshooter; Lake Image Systems Ltd; Tring, England) at a rate of 250 frames/s (high speed video microscopy) or 30 frames/s (standard video microscopy) as previously described.9 Usually a frame rate of 500 frames/s and × 100 objective is recommended; however, under these conditions, a higher light intensity is required to obtain an acceptable-quality image. In our study, the higher light intensity prevented the sample from being effectively cooled due to heat transference from the bulb and oil immersion lens. Recordings were made at the following temperatures: 37°C, 32°C, 27°C, 22°C, 17°C, 12°C, 7°C, and 2°C. Intermediary temperatures also were recorded when possible to help to construct a more detailed model. For each recording, five readings of CBF were taken from different areas along each ciliated edge. These readings were converted to CBF by a simple calculation [CBF = frame rate/(number frames for five beats) × 5]. Ciliary dyskinesia and beat amplitude were scored as described previously.11,12 Briefly, dyskinesia was defined as an abnormal beat pattern that included reduced beat amplitude, stiff beat pattern, failure to bend along the length of the ciliary shaft, flickering or a twitching motion, and static cilia.