Anti cd3 clone cd3 12

Skin, lung, liver, colon, small intestine, stomach, lacrimal gland, and salivary gland samples were fixed with 4% paraformaldehyde and imbedded in paraffin. Sections (5 μm) were stained with hematoxylin and eosin. Skin sections were also stained with anti-CD3 (clone CD3-12; Serotec). Images were acquired with a BX51 discussion microscope (Olympus) with PixeLink camera under 100× magnification.

Granulomas were excised from the dorsal skin region one week after the third injection (i.e., 13 weeks of age) and fixed in 10% formalin solution overnight at room temperature. Fixed tissues were processed to paraffin blocks by the Molecular Pathology Core at the University of Florida. Paraffin sections (4 μm) were stained with H&E, or by immunohistochemistry with the following primary antibodies: anti- CD45R/B220, clone RA3-6B2 (BD Biosciences, San Jose, CA, USA), anti- CD3, clone CD3-12 (Serotec, Raleigh, NC, USA), anti- F4/80, clone BM8 (Caltag Medsystems, Buckingham, UK), or anti- LYVE-1 (Lymphatic vessel endothelial hyaluronan receptor-1; Abcam, Cambridge, MA, USA). Deparaffinized sections were hydrated then blocked in normal goat serum followed by incubation in primary antibodies for one hour at room temperature. Following washes, sections were incubated with goat anti-rabbit HRP-polymer labeled antibody (Mach2 HRP, Biocare, Concord, CA, USA) for 30 min at room temperature followed by horseradish peroxidase- DAB and hematoxylin counterstain.