Intestinal Permeability Quantification

For intestinal permeability status, immunofluorescence using an anti-Occludin antibody (Abcam, Cambridge, UK) was used in paraffin cut sections of jejunum. A Cy3-labeled secondary antibody was used and slides were mounted in a DAPI-containing solution (Vector Laboratories, Burlingame, CA, USA). Three images per section were taken using a confocal immunofluorescent microscope, Zeiss LSM 800 Axio Observer (Zeiss, Thornwood, NY, USA) (magnification 40×), and fluorescence intensity was analyzed with Image J software. Fluorescence measured at the lumen of the colonic tissue was used as background fluorescence to subtract from the epithelial fluorescence values. Therefore, fluorescence density values represented the total protein in the epithelial cells [78 (link)].