Analysis and isolation of peritrophic membrane in I. scapularis.

(A) Transmission electron microscopic detection of peritrophic membrane (PM). While undetectable in the gut of unfed ticks (left panel), a PM structure is conspicuous in ticks and was isolated and processed at 36 h of feeding on mice (L, lumen; EC, gut epithelial cell). Arrows point to the PM. Scale bar, 5 μm. (B) Immunofluorescence localization of PM. Cryosections from unfixed tick gut samples were labeled with WGA-FITC (green) and propidium iodide (PI, red) and imaged using a confocal microscope. Arrows point to the green PM, while the nuclei (N) of gut epithelial cells are labeled red. Scale bar, 10 μm. (C) Isolation of an intact PM structure. The PM was isolated from the dissected gut of nymphal ticks and viewed under a binocular dissecting microscope. Scale bar, 100 μm. (D) Relative purity of the extracted PM. The PM, as shown in Figure 1C, was stained with WGA-FITC (green) and PI (red) and imaged using a confocal microscope. Lack of red fluorescence indicated absence of contaminating gut cells. Scale bar, 20 μm.