By William R. Skach
Cystic Fibrosis: tools and Protocols consolidates state of the art in vitro, mobile, and entire animal laboratory protocols into an necessary source. From electrophysiology and mobile biology, to animal versions and gene treatment, this accomplished set of tools presents the step by step directions wanted for investigators to include new methods into their examine courses. particular protocols describe new innovations for prognosis, in vitro tools for the expression and useful research of CFTR, novel biochemical and mobile structures to figure out how mutations subvert CFTR functionality, and in vivo protocols to envision how CFTR disorder produce multisystem pathology in either human and animal types.
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Extra info for Cystic Fibrosis Methods and Protocols
Identification of CFTR Using Physiological Characteristics The initial task during a recording is to identify CFTR as the current carrier. Conditions must be used that immediately identify the ion species that carries the measured current. This is most simply done by excluding most other small ions in the bath and pipet filling solution so that Cl– is the major current-carrying ion. Commonly, small cations are exchanged for the large cation N-methylD-glucamine (NMDG), which does not support ion currents through most biological channels.
Mol. Cell. Biol. 11, 3886–3893. 20. Cheng, S. , Gregory, R. , Souza, D. , White, G. , et al. (1990) Defective intracellular transport and processing of CFTR is the molecular basis of most cystic fibrosis. Cell 63, 827–834. 21. Ward, C. , and Kopito, R. R. (1995) Degradation of CFTR by the ubiquitin-proteasome pathway. Cell 83, 121–127. 22. Welsh, M. J. and Smith, A. E. (1993) Molecular mechanisms of CFTR chloride channel dysfunction in cystic fibrosis. Cell 73, 1251–1254. 23. Choi, J. , Lee, M.
6. Technique for Acquiring Patch Clamp and Noise Data Analog electrophysiological signals have to be appropriately digitized for subsequent analysis. Patch clamp analysis and noise analysis require significantly different methods of digitizing, although the analog signal for both analyses is initially the same. Single-channel patch clamp data are generally sampled at a sampling frequency (fs) that is five times the filter frequency (ff). 5 (for an ideal filter, fs/ff = 2, which is the Nyquist rate).