Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up
Fluorescently labelled compounds are often employed to study the paracellular properties of epithelia. For flux measurements, these compounds are added to the donor compartment and samples collected from the acceptor compartment at regular intervals. However, this method fails to detect rapid change...
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| Format: | Article |
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MDPI AG
2024-02-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/25/4/2252 |
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| author | Çlirim Alija Lukas Knobe Ioanna Pouyiourou Mikio Furuse Rita Rosenthal Dorothee Günzel |
| author_facet | Çlirim Alija Lukas Knobe Ioanna Pouyiourou Mikio Furuse Rita Rosenthal Dorothee Günzel |
| author_sort | Çlirim Alija |
| collection | DOAJ |
| description | Fluorescently labelled compounds are often employed to study the paracellular properties of epithelia. For flux measurements, these compounds are added to the donor compartment and samples collected from the acceptor compartment at regular intervals. However, this method fails to detect rapid changes in permeability. For continuous transepithelial flux measurements in an Ussing chamber setting, a device was developed, consisting of a flow-through chamber with an attached LED, optical filter, and photodiode, all encased in a light-impermeable container. The photodiode output was amplified and recorded. Calibration with defined fluorescein concentration (range of 1 nM to 150 nM) resulted in a linear output. As proof of principle, flux measurements were performed on various cell lines. The results confirmed a linear dependence of the flux on the fluorescein concentration in the donor compartment. Flux depended on paracellular barrier function (expression of specific tight junction proteins, and EGTA application to induce barrier loss), whereas activation of transcellular chloride secretion had no effect on fluorescein flux. Manipulation of the lateral space by osmotic changes in the perfusion solution also affected transepithelial fluorescein flux. In summary, this device allows a continuous recording of transepithelial flux of fluorescent compounds in parallel with the electrical parameters recorded by the Ussing chamber. |
| first_indexed | 2024-03-07T22:28:50Z |
| format | Article |
| id | doaj.art-a70150b844b44f7d9e521cfa985a2011 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-07T22:28:50Z |
| publishDate | 2024-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-a70150b844b44f7d9e521cfa985a20112024-02-23T15:20:20ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672024-02-01254225210.3390/ijms25042252Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-UpÇlirim Alija0Lukas Knobe1Ioanna Pouyiourou2Mikio Furuse3Rita Rosenthal4Dorothee Günzel5Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité–Universitätsmedizin Berlin, 12203 Berlin, GermanyClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité–Universitätsmedizin Berlin, 12203 Berlin, GermanyClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité–Universitätsmedizin Berlin, 12203 Berlin, GermanyDivision of Cell Structure, National Institute for Physiological Sciences, Okazaki, Aichi 444-8787, JapanClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité–Universitätsmedizin Berlin, 12203 Berlin, GermanyClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité–Universitätsmedizin Berlin, 12203 Berlin, GermanyFluorescently labelled compounds are often employed to study the paracellular properties of epithelia. For flux measurements, these compounds are added to the donor compartment and samples collected from the acceptor compartment at regular intervals. However, this method fails to detect rapid changes in permeability. For continuous transepithelial flux measurements in an Ussing chamber setting, a device was developed, consisting of a flow-through chamber with an attached LED, optical filter, and photodiode, all encased in a light-impermeable container. The photodiode output was amplified and recorded. Calibration with defined fluorescein concentration (range of 1 nM to 150 nM) resulted in a linear output. As proof of principle, flux measurements were performed on various cell lines. The results confirmed a linear dependence of the flux on the fluorescein concentration in the donor compartment. Flux depended on paracellular barrier function (expression of specific tight junction proteins, and EGTA application to induce barrier loss), whereas activation of transcellular chloride secretion had no effect on fluorescein flux. Manipulation of the lateral space by osmotic changes in the perfusion solution also affected transepithelial fluorescein flux. In summary, this device allows a continuous recording of transepithelial flux of fluorescent compounds in parallel with the electrical parameters recorded by the Ussing chamber.https://www.mdpi.com/1422-0067/25/4/2252transepithelial fluxparacellular fluorescent markerUssing chamberautomatized flux measurementtight junctionosmotic stress |
| spellingShingle | Çlirim Alija Lukas Knobe Ioanna Pouyiourou Mikio Furuse Rita Rosenthal Dorothee Günzel Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up International Journal of Molecular Sciences transepithelial flux paracellular fluorescent marker Ussing chamber automatized flux measurement tight junction osmotic stress |
| title | Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up |
| title_full | Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up |
| title_fullStr | Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up |
| title_full_unstemmed | Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up |
| title_short | Integrating Continuous Transepithelial Flux Measurements into an Ussing Chamber Set-Up |
| title_sort | integrating continuous transepithelial flux measurements into an ussing chamber set up |
| topic | transepithelial flux paracellular fluorescent marker Ussing chamber automatized flux measurement tight junction osmotic stress |
| url | https://www.mdpi.com/1422-0067/25/4/2252 |
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