Methacarn-fixed sections from the colons of WT and 1KO mice were stained by immunohistochemistry for the indicated markers. (A) 40X fluorescent microscopy images of colon tissue stained for MUC2 (peptide sequence of mucin), Alcian blue (AB)(acidic mucins), Mucicarmine (acidic mucins) and Periodic Acid Schiff and Alcian Blue (PAS/AB) (neutral/acidic mucins). Scale bar equals 50 microns. Confocal images of GOB5 (mCLCA3) staining of the distal colon. (B) Alcian blue stained distal colonic sections from WT and 1KO mice were used for quantification of (C) mucus layer thickness and (D) Alcian blue positive vesicles per crypt. (E) Relative ROS activity in mucosa and colonic contents. (F) 10X and 40X fluorescent microscope images of distal colon stained with antibodies against lysozyme and a-defensin-1 (DEFA-1) and 10X image of isotype control. Scale bar equals 50 microns. A–B and F, representative images taken from 3–5 mice per group; C and D, individual measurements of mucus thickness along the distal colon taken from 3 mice per group; E, each point is an individual mouse from two different experiments. C and D, ****, p<0.0001 Mann-Whitney Test; F, *, p = 0.02 Students’ t-test corrected for multiple comparisons (Holm-Sidak