Figure 7. High affinity LFA-1 is not... | Rockefeller University Press

Figure 7.

$Figure 7. High affinity LFA-1 is not required for uropodal adhesion. (A and B) Human primary T lymphocytes were pretreated with blebbistatin and incubated on cover glasses coated with ICAM-1 and CXCL-12 for 20 min. Cells were then processed for dual immunofluorescent labeling with TS2/4 antibody and KIM127 (A) or m24 (B) antibody and subjected to TIRF microscopy. Ratio images were generated by subtracting the background and dividing KIM127 (A) or m24 (B) intensity by TS2/4 intensity. Line profiles of LFA-1 intensity are presented along the front (Head)-to-back (Tail) line (A). (B) From three independent experiments, a total of 48 (KIM127/TS2/4) and 35 (m24/TS2/4) cells was randomly selected, carefully analyzed, and scored for the presence of KIM127 or m24 staining enriched at the anterior region, the posterior region, or both, based on the ratio images. Each bar represents the percentage of total cells scored. The black portion of each bar is the fraction of cells scored for the anterior region dominant staining of KIM127 or m24. The white portion of each bar is the fraction of the cells that showed even distribution of staining. The gray portion of each bar is the fraction of cells scored for the posterior region dominant staining of KIM127 or m24. Bar, 20 μm.$

High affinity LFA-1 is not required for uropodal adhesion. (A and B) Human primary T lymphocytes were pretreated with blebbistatin and incubated on cover glasses coated with ICAM-1 and CXCL-12 for 20 min. Cells were then processed for dual immunofluorescent labeling with TS2/4 antibody and KIM127 (A) or m24 (B) antibody and subjected to TIRF microscopy. Ratio images were generated by subtracting the background and dividing KIM127 (A) or m24 (B) intensity by TS2/4 intensity. Line profiles of LFA-1 intensity are presented along the front (Head)-to-back (Tail) line (A). (B) From three independent experiments, a total of 48 (KIM127/TS2/4) and 35 (m24/TS2/4) cells was randomly selected, carefully analyzed, and scored for the presence of KIM127 or m24 staining enriched at the anterior region, the posterior region, or both, based on the ratio images. Each bar represents the percentage of total cells scored. The black portion of each bar is the fraction of cells scored for the anterior region dominant staining of KIM127 or m24. The white portion of each bar is the fraction of the cells that showed even distribution of staining. The gray portion of each bar is the fraction of cells scored for the posterior region dominant staining of KIM127 or m24. Bar, 20 μm.