Whole frozen sections were processed for indirect immunofluorescence and confocal microscopy as described [12 (link)], which were prepared from paraformaldehyde-fixed tissues. Briefly, 10-μm frozen sections of proximal colon samples were collected on coated slides. Then it was washed three times with PBS and blocked with 5% bovine serum albumin (sigma Aldrich) at room temperature for 30 min. After staining using the primary antibodies, rabbit anti-mouse CgA (1:500; Abcam), rat anti-mouse 5-HT (1:50; Abcam), and secondary antibodies conjugated to Alexa fluor 488 or 594 (Molecular Probes). Images were examined using a confocal microscope (Olympus, Germany).
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