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Recruitment of macrophages into CNV lesions over time was evaluated by FACS in single-cell suspensions originating from mouse retinal tissue.
In mouse retina, more than 50 to 100 cells/retina seemed morphologically intact, but 17.3% of the mouse retinal tissue after organotypic culture showed less viability (Figure 3E, YO-PRO-1 staining).
Indeed, we observed decreases in CF3 fluorescence upon acute bath application of the copper chelator BCS to dissociated hippocampal neurons as well as isolated mouse retinal tissue, whereas Ctrl-CF3 fluorescence remained unchanged by this treatment.
Western blotting on mouse retinal tissue lysates revealed that the healthy, murine retina contains very low amounts of MMP-9, adding to the difficulties to accurately discern MMP-9 on Western blot.
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The SAGE data were generated from mouse retinal tissues at 10 different developmental stages ranging from E12.5 (Theiler stage 20) to post natal day 10 (P10) and adult; the data were originally analyzed using the PoissonC algorithm with K = 24.
These results suggested that activated Lyn is not detectable by western blot in mouse retinal tissues.
We observed differences in expression patterns of SFK members Fyn and Lyn in RF/6A retinal endothelial cells versus mouse retinal tissues, with c-Src predominating in retinal tissues.
Moreover, western blot analysis using an antibody against Fyn revealed no detectable expression of Fyn in mouse retinal tissues as compared to RF/6A cells (Fig. 7B).
Quantitation of the immunostaining in retinal blood vessels revealed a 10-fold10-foldse in activated SFK levels in sections of mouse retinal tissues in which Tbdn endothelial expression was knocked down for 6-week compared to control (P<0.00001; Fig. 6B).
To confirm our immunohistochemical data indicating that blood vessels of retinal lesions of endothelial Tbdn knockdown mice are associated with an up-regulation of levels of activated SFK, western blot analyses with phospho-Src family (Tyr416) antibody were performed on isolated mouse retinal tissues in which Tbdn endothelial expression was knocked down for 6-weeks versus control.
To determine what molecular factors might be associated with Chrnb2 mutation at the age when cholinergic waves are normally present (P1 P8), we assayed RNA populations in both mutant and WT (C57BL/6J) mouse LGN and retinal tissue at P4 by microarray hybridizations.
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