Leukocyte Migration In Vivo Figure 8. Migration patterns of GFP-positive and GFP-negative cells. A: representative in vivo microscopy images show transmigration of GFP-positive (GFP-image) and GFP-negative cells (RLOT-image) at 60 min after microinjection of MCP-1 in Cx3CR1gfp/gfp mice (objective magnification 206; asterisk shows the site of microinjection). B–E: The motility parameters, curve-line migration velocity (B), straight-line migration velocity (C), and directionality (D) of interstitially migrating, GFP-positive (monocytes, gray bars) and GFP-negative cells (neutrophils, black bars) were analyzed at 60, 70, and 80 min after microinjection of MCP-1 for 5 min, respectively; mean6SEM; *p,0.05 vs. GFP-negative cells; # p,0.05 vs. 60–65 min; n = 5. E: polarization of GFP-negative cells (black bars) and GFP-positive cells (gray bars) analyzed at 70 min after microinjection of MCP-1; mean6SEM, n = 15. doi:10.1371/journal.pone.0004693.g008 adhesion, transmigration, and motility of transmigrated leukocytes as compared to microinjection of saline. Second, we compared the character of leukocyte migration after microinjections of chemoat- tractants or E. coli with that induced by intrascrotal injection. Leukocyte adhesion, transmigration, and interstitial migration were evaluated on the vessel side ipsilateral to the microinjection site and compared with those on the contralateral side. As a result, intrascrotal injection of PAF induced diffuse tissue inflammation without any prevalence of leukocyte adhesion or transmigration on either the ipsi- or the contralateral side. Likewise, leukocyte interstitial migration upon intrascrotal injection of PAF displayed a diffuse character as shown by low levels of leukocyte straight-line PLoS ONE | www.plosone.org 8 March 2009 | Volume 4 | Issue 3 | e4693