Why does null hypothesis testing matter for FITC-dextran permeability assays?

Null hypothesis testing enables objective determination of whether observed differences in intestinal fluorescence reflect true permeability changes or random variation, supporting rigorous target validation.

How does independent variable isolation fit the FITC-dextran workflow?

Isolating variables such as bacterial strain or compound exposure ensures that changes in fluorescence are attributable to the intervention, strengthening mechanistic insights in the discovery pipeline.

What do quantitative dependent variable measurements enable in this assay?

Quantitative fluorescence measurements allow precise comparison of intestinal permeability across experimental groups, enabling robust statistical analysis and data-driven decision making.

Why are replication requirements critical for cross-functional collaboration in permeability studies?

Replication ensures that permeability findings are reproducible and reliable, facilitating data sharing and alignment across discovery, screening, and translational teams.

What statistical analysis capabilities are required before implementing FITC-dextran readouts?

Teams must be equipped to perform statistical comparisons of fluorescence intensity, account for background autofluorescence, and validate significance thresholds to support actionable R&D decisions.

FITC-Dextran Feeding: A Method to Quantify Intestinal Permeability in C. elegans

JoVE Encyclopedia of Experiments

Biology

0 views • 2:56 min • April 30th, 2023

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- C. elegans is used as an experimental model of infection by intestinal pathogens which cause disease, in part, by disrupting the intestinal barrier, increasing its permeability. To quantify barrier integrity after exposure to a bacteria or chemical compound of interest, collect worms by washing them off their plates.

Next, place these worms on agar plates with labeled food composed of heat-killed E. coli mixed with FITC-dextran, a high molecular weight polysaccharide with a fluorescent tag.

Allow the worms to eat the labeled bacteria. Those with intact intestinal barriers will rapidly clear the food, while in worms with more permeable intestines, the fluorescent marker will penetrate beyond the lumen of the organ.

Transfer worms to a formaldehyde solution to fix the animals. After 1 to 2 minutes, replace the formaldehyde with mounting media.

Finally, image the worms by fluorescent microscopy. Note that C. elegans intestinal granules are autofluorescent, so there will be background signal. But overall, more intestinal fluorescence indicates higher permeability.

In the example protocol, we will assay the effect of treatment with the biomolecule DIM on the intestinal permeability of worms infected with pathogenic Pseudomonas aeruginosa bacteria.

- Prepare the FITC-dextran-supplemented plates by mixing 2 milliliters of heat-inactivated E. coli with 4 milligrams of FITC-dextran. Add 100 microliters of the mixture to each of 20 fresh NGM agar plates and allow the plates to dry for one hour on a clean lab bench. After 48 hours of treatment, wash the worms with S Buffer. Transfer them to FITC-dextran supplemented plates and to NGM plates without FITC-dextran and incubate the plates overnight.

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Last updated: 25 July 2026

FITC-Dextran Feeding: A Method to Quantify Intestinal Permeability in C. elegans

Overview

Key Study Components

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Purpose of Study

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Frequently Asked Questions

Executive Industry Relevance

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Early Discovery & Target Validation

Screening & Assay Development

Translational & Preclinical Research

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