Quantification of Glandular Inflammation

Quantitation of lacrimal and salivary gland inflammation was performed as previously described [53 (link)]. Briefly, exorbital lacrimal glands and submandibular salivary glands were fixed in formalin, processed, embedded in paraffin, and 5 µm sections were stained with hematoxylin and eosin (H&E). Inflammation was quantified in a blinded manner by standard light microscopy at 10× objective using standard focus-scoring with a focus defined as an aggregate of at least 50 mononuclear cells and the focus score defined as the number of foci per 4 mm² of tissue. Tissue areas were calculated by ImageJ software [54 (link)] using low magnification digital images obtained by scanning H&E-stained sections with the PathScan Enabler IV (Meyer Instruments, Houston, TX, USA). The WT lacrimal and salivary glands in Figure 1 were previously published in comparison to another KO NOD strain [21 (link)] from the same colony as the Tlr7 KO samples to which they were compared in this study. Representative H&E-stained sections in the figure were obtained by whole-slide scans using the PathScan Enabler 5 (Meyer Instruments).