Intestinal permeability measurement was performed as previously described [49 (link)]. Following mice euthanasia, jejunal fragments were immediately mounted in Ussing chambers (Physiologic Instruments) and incubated 2 h with Kreb's solution (Sigma) constantly oxygenated with carbogen (95% O2, 5% CO2) in presence of both Fluorescein Sodium Salt (FSS 376 Da, Sigma) and Horse Radish Peroxidase (HRP 4 kDA; Sigma) in the mucosal compartment. Gut integrity was continuously checked through electro-physiological measurements and electrical resistance was monitored (R, Ω.cm2). After 2h of incubation, the serosal compartment was sampled to follow the epithelial paracellular passage of FSS by measuring the fluorescence intensity (485nm/525 nm) using a microplate reader (Spark®). Epithelial permeability to HRP was determined by ELISA as previously described [49 (link)].
Fluorescein sodium salt fss 376 da
Manufactured by Merck Group
Sourced in France
Fluorescein Sodium Salt (FSS) is a laboratory reagent with a molecular weight of 376 Da. It is a yellow-green crystalline powder that is highly soluble in water. FSS is commonly used as a fluorescent dye in various analytical and research applications.
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2 protocols using fluorescein sodium salt fss 376 da
1
Intestinal Permeability Measurement Protocol
2
Evaluating Intestinal Permeability in Rat Colonic Tissue
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Colonic fragments from rats fed control or DON-contaminated diets were mounted in Ussing chambers (Easymount, Physiologic Instruments, Hamden, USA) exposing a surface area of 0.1cm 2 (Riba et al., 2018) (link). Oxygenated Krebs's solution (Sigma, Saint-Quentin Fallavier, France) thermostated at 37 °C was put on each side. Electrical resistance (R) was recorded over the 2-h period of the experiment. Fluorescein sodium salt (FSS, 376 Da, Sigma Saint-Quentin Fallavier, France) used as a marker of paracellular permeability and horseradish peroxidase (HRP, 44 kDa, Sigma Saint-Quentin Fallavier, France) used as a marker of transcellular permeability were added in the mucosal compartment at 40 µg/mL and 0.4 mg/mL concentrations, respectively. Epithelial permeability to FSS was determined by measuring fluorescence intensity at 485 nm/525 nm using an automatic Infinite M20 microplate reader (Tecan, Austria). Epithelial permeability to intact HRP was determined as previously described (Payros et al., 2014) (link), by an enzymatic assay for the specific HRP activities found in the serosal and mucosal compartments, using the same Infinite M20 microplate reader. Permeability was calculated as the ratio of flux to the initial concentration and expressed in cm/second.
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