Figure 2: Identification of RANKL using immunofluorescence. (a) Representative micrographs illustrate immunofluorescence of antibodies against RANKL at 4-day control and 8- and 15-day ligated groups. Note RANKL expression around the vasculature (V) and endosteal spaces (asterisks). Multinucleated osteoclast-like cells were also observed at the PDL-bone interface (white arrows). (b) Higher magnification micrographs show RANKL immunofluorescence in local regions of the complex at 15 days of ligation. (c) Schematic of rat periodontal tissue (mesiodistal section) with gray boxes (125 μm × 125 μm) that indicate target areas used to count RANKL(+) cells. (d, e) Bar graphs illustrate RANKL(+) cell count within specified target areas between control and ligated groups on mesial (d) and distal (e) sides. *Statistically significant difference at 95% confidence interval was observed. Junctional epithelium (JE), epithelium (E), periodontal ligament (PDL), alveolar bone (AB), alveolar bone crest (ABC), dentin (D), and secondary cementum (SC).