Cftr Assay
Cftr Assay - Related Videos
10:05
In Vitro Analysis of PDZ-dependent CFTR Macromolecular Signaling Complexes
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Cited by 7 •
2012
Cystic fibrosis transmembrane conductance regulator (CFTR), an epithelial chloride channel, has been reported to interact with various proteins and regulate important cellular processes; among them the CFTR PDZ motif-mediated interactions have been well documented. This protocol describes methods we developed to assemble a PDZ-dependent CFTR macromolecular signaling complex in vitro.
09:59
Functional Reconstitution and Channel Activity Measurements of Purified Wildtype and Mutant CFTR Protein
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Cited by 7 •
2015
Described here is a rapid and effective procedure for functional reconstitution of purified wild-type and mutant CFTR protein that preserves activity for this chloride channel, which is defective in Cystic Fibrosis. Iodide efflux from reconstituted proteoliposomes mediated by CFTR allows studies of channel activity and the effects of small molecules.
09:13
Cell-surface Biotinylation Assay
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2023
A cell can regulate the amount of particular proteins on its cell membrane through endocytosis, following which cell surface proteins are effectively sequestered in the cytoplasm. Once within a cell, these surface proteins can be either destroyed or “recycled” back to the membrane. The cell surface biotinylation assay provides researchers with a way to study these phenomena. The technique makes use of a derivative of the small molecule biotin, which can label surface proteins and then be...
07:04
Forskolin-induced Swelling in Intestinal Organoids: An In Vitro Assay for Assessing Drug Response in Cystic Fibrosis Patients
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Cited by 121 •
2017
This protocol describes an assay for measuring CFTR function and CFTR modulator responses in cultured tissue from subjects with cystic fibrosis (CF). Biopsy-derived intestinal organoids swell in a cAMP-driven fashion, a response that is defective (or strongly reduced) in CF organoids and can be restored by exposure to CFTR modulators.
08:12
The TUNEL Assay
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2023
One of the hallmarks of apoptosis is the nuclear DNA fragmentation by nucleases. These enzymes are activated by caspases, the family of proteins that execute the cell death program. TUNEL assay is a method that takes advantage of this feature to detect apoptotic cells. In this assay, an enzyme called terminal deoxynucleotidyl transferase catalyzes the addition of dUTP nucleotides to the free 3’ ends of fragmented DNA. By using dUTPs that are labeled with chemical tags that can produce...
FAQs
cAMP stimulation activates CFTR in cells, producing chloride movement across the membrane. The assay then measures the resulting response with an electrical, fluorescent, or other ion-sensitive readout. The size of this response provides functional evidence about channel activity, allowing researchers to evaluate whether CFTR can respond appropriately under the test conditions.
These measurements represent different biological problems. Reduced abundance indicates that less CFTR protein is present, while abnormal localization suggests defective trafficking to the relevant membrane. Normal abundance and localization can still coexist with impaired channel activity. Separating these outcomes helps researchers characterize disease-associated defects more precisely rather than treating every abnormal result as the same mechanism.
Electrical, fluorescent, and other ion-sensitive readouts provide different ways to quantify the chloride response after stimulation. Their shared purpose is to convert ion movement into a measurable assay signal, while the selected format determines how the response is detected. This flexibility allows investigators to match the readout with the cellular question, whether it concerns channel function or treatment response.
A basic workflow begins by choosing whether to measure CFTR abundance, localization, function, or a combination of these properties. Cells are then evaluated under defined conditions, including cAMP stimulation when functional activity is the target, and the response is quantified with an appropriate readout. Comparing these measurements helps separate expression or trafficking defects from impaired channel performance.
In medicine and disease research, assay results can show whether a variant is associated with reduced CFTR abundance, abnormal localization, impaired channel activity, or a combination of defects. This functional information supports classification of disease-associated variants and adds biological context beyond identifying a genetic change alone. The findings can therefore contribute to understanding how specific variants affect epithelial fluid balance.
Researchers can measure CFTR function before and after exposure to a modulator to determine whether channel activity improves. Depending on the assay design, they may also examine whether treatment affects protein abundance or localization. These outcomes indicate whether a compound restores functional performance and help support precision treatment approaches for cystic fibrosis by connecting cellular responses with individual CFTR defects.