Immunofluorescence labeling of apical and basolateral proteins demonstrated reorganization of the membrane proteins into a polarized distribution during spheroid culture.
As observed with the co-expression of PIP5K4, the association of SYP with the apical plasma membrane increased and the typical higher labelling of apical vesicles dissipated (figure 6A,C).
NF2 IHC in normal ependymal lining and ependymomas showed only faint or no labeling of apical PM, respectively, suggesting that NF2, unlike moesin, is not a major ligand of NHERF1 in these polarized structures.
Accompanying the partial loss of polarity, the association of GFP fluorescence with the apical plasma membrane increased, the typical higher labelling of the apical cytoplasm dissipated (figure 5A-C), and some cells (n = 4) exhibited large labelled structures in the cytoplasm that extended to the apical region (figure 5D).
When FM1-43 wappliedind in the presence of 5μM NMB-1, the increase in cytoplasmic fluorescence decreased by 80 85% in IHCs and OHCs (Fig. 7a c) and displayed a transient temporal profile associated with non-specific labeling of the apical plasma membranes of these cells (Fig. 7b).
In glandular epithelia, among them colonic enterocytes, mAb 22-1-1 22-1-1 22-1-1nular labexhibitedthe apical portion of cells.
Within tracheal ciliated cells, a more intense labelling of the apical cytoplasmatic region was noted, consistent with an earlier report on human bronchi (Klapproth et al. 1997).
A third example with striking differences of staining patterns between both antibodies includes normal prostate tissue where staining with 22-1-1 22-1-1 22-1-1landular epithelia wash a prominent labeling of the apical cytoplaseen portinns.
To record the distribution of label from apical to basal portions of the cell, z-series were captured of the cells at 0.5 micron steps.
This could be seen in sagittal views (Fig. 7A,C) and by quantitation of apical and lateral labeling intensities (Fig. 7B,D).
