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Table 2 Advantages and limitations of visual and imaging methods and techniques used for measuring mucus properties

From: Strategies for measuring airway mucus and mucins

Beads/microspheres visualization and tracking in vitro [87,88,89], ex vivo [20, 90] and in vivo [91]- Easily visualized.
- Potential for in vivo tracking.
- Most applicable in vitro and in small animal models ex vivo/in situ.
- Data analysis can require careful application of modelled calculations that present opportunities for error.
- Some of the in vivo applications may require expensive visualization set up.
Histology & Immunostaining (using specific antibodies, Lectins, PAS/AB) [90, 92, 93]- Inexpensive, easily visualized.
- Specific antibodies can provide precise mucin detection and localization or co-localization with other molecules.
- Fluorescent lectins can be used for semi-quantitation by fluorescence intensity measurement and are inexpensive
- Applicable mostly in vitro and ex vivo.
- When scoring systems are utilized, careful analysis by multiple individuals blinded to group treatments are necessary.
- Fixation and washing steps might result in mucus being washed away.
- Lectins bind to different carbohydrates in the oligosaccharide chains of glycoproteins and glycolipids and therefore are not mucin specific.
Electron microscopy [94]- In depth view of micro anatomical structures of cells and gel-forming mucins.- Difficult to detect more than one type of gel mucin at the same time.
- The type and duration of fixation is very important for retention of mucin structures.
X-ray imaging analysis [1, 20, 95,96,97,98]- Novel techniques provide in vivo ability to detect mucus
- Very recent X-ray synchrotron [96] and quazi-monochromatic X-ray phase-contrast imaging techniques have been applied successfully to measure MCT in vivo, together with lung motion.
- Can detect mucus plugs in humans in vivo.
- Expensive set-up and materials.
- Potential for exposure to harmful rays.
- At the moment, are limited in utility for longitudinal in vivo studies.
- Highly specialized equipment and skills
Volumetric – submucosal gland bubble visualization [55, 99]- Detect ex vivo/in vitro increased output from single cell or multiple glands under normal or treatment conditions.
- The total volume technique gives a simple quantitation of total mucus secretion ex vivo/in vitro for a constant time period at baseline and/or after treatment.
- Volume output may not necessarily comprise only mucus but can also include changes in serous gland- and non-glandular cell-secretions.
- Currently not applicable in vivo.